Synthesis and cell phototoxicity of a triply bridged fused diporphyrin appended with six thioglucose units

Article abstract of DOI:10.1016/j.tetlet.2014.09.085

A triply bridged fused diporphyrin appended with six thioglucose units is reported. This new, chemically, and photochemically stable amphiphilic compound is taken up by breast cancer cells and causes cell death upon light exposure. Photophysical studies r

Full text of DOI:10.1016/j.tetlet.2014.09.085

Tetrahedron Letters 55 (2014) 6311–6314
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Tetrahedron Letters
journal homepage: www.elsevier.com/locate/tetlet
Synthesis and cell phototoxicity of a triply bridged fused diporphyrin
appended with six thioglucose units
Sunaina Singh a,b, , Amit Aggarwal a,b, , N. V. S. Dinesh K. Bhupathiraju a, Brandon Newton c,
Ahmad Nafees b, Ruomei Gao c,d, Charles Michael Drain a,e,
⇑
a Department of Chemistry & Biochemistry, Hunter College of The City University of New York, 695 Park Avenue, New York, NY 10065, USA
b Department of Natural Sciences, LaGuardia Community College of The City University of New York, 31-10 Thomson Avenue, Long Island City, New York, NY 11101, USA
c Department of Chemistry and Biochemistry, Jackson State University, Jackson, MS 39211, USA
d Chemistry and Physics Department, SUNY College at Old Westbury, Old Westbury, NY 11568, USA
e The Rockefeller University, 1230 York Avenue, New York, NY 10065, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 23 July 2014
Revised 16 September 2014
Accepted 18 September 2014
Available online 28 September 2014
A triply bridged fused diporphyrin appended with six thioglucose units is reported. This new, chemically,
and photochemically stable amphiphilic compound is taken up by breast cancer cells and causes cell
death upon light exposure. Photophysical studies reveal absorption bands in the near IR region, and
photosensitized formation of singlet oxygen in high quantum yields.
Ó 2014 Elsevier Ltd. All rights reserved.
Keywords:
Porphyrin
Thioglucose
Photodynamic therapy
Fluorescence
Click chemistry
Introduction
is a need for a palate of these core dye platforms with different
photophysical properties. Porphyrinoids are generally non-toxic
There are many applications of dyes in biochemistry, imaging,
and therapy that require appended, robust biotargeting motifs.
Since there is a wide range of biotargeting motifs available for all
of these applications, core dye platforms with appropriate photo-
physical properties that can be rapidly and efficiently appended
with the targeting motif avoid the complex redesign of synthetic
strategies. Photodynamic therapy (PDT), for example, is a non-
invasive treatment for cancer involving the interaction of light, a
photosensitizer and oxygen to result in generation of singlet oxy-
gen and other reactive oxygen species that cause necrosis or apop-
tosis.1,2 There are many cancer targeting motifs, including sugars
and peptides.
The photophysics of porphyrin and phthalocyanine derivatives
can be tuned for many of the above applications. For example,
those with good triplet quantum yields can serve as photosensitiz-
ers for PDT, while those with good fluorescence quantum yields
can serve as imaging agents and biochemical trackers. Thus there
under dark conditions.3,4 Imaging and PDT of various forms of can-
cers are more efficient if the dye absorbs light in the therapeutic
window (ca. 700–1100 nm) especially for deep cancers,5–7 so bio-
targeted dye systems with longer wavelength absorptions are
needed for next-generation imaging and PDT agents.8,9
Direct, covalent linking of porphyrins (e.g., 2 and 3 in Fig. 1)
yields systems with photophysical properties such as high polariz-
ability and high nonlinear optical character10 that arise from strong
electronic coupling of the macrocycles. Compounds such as 1 and 2
are fluorescent. Fused (b–b, meso–meso, b–b) triply bridged por-
phyrins have low energy absorption bands in the infrared region
because of extended conjugation and coplanar geometries.11 Com-
pounds such as 3 are minimally fluorescent but exhibit large two-
photon absorption (2PA) cross sections (r) where two low energy
1400–2000 nm photons are simultaneously absorbed and have
good triplet quantum yields.12,13 Sugar moieties appended to por-
phyrinoids increase the selective uptake by cancer cells.14 The
number, type, and position of the sugar moieties effect cell
uptake,15,16 but the hydrolysis of O-glyco linkages of many
reported conjugates can diminish in vivo effectiveness compared
to non-hydrolysable derivatives.14
⇑
Corresponding author. Fax: +1 212 7725332.
E-mail address: cdrain@hunter.cuny.edu (C.M. Drain).
S.S. and A.A. contributed equally to this work.
http://dx.doi.org/10.1016/j.tetlet.2014.09.085
0040-4039/Ó 2014 Elsevier Ltd. All rights reserved.

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S. Singh et al. / Tetrahedron Letters 55 (2014) 6311–6314
Figure 1. Oxidative coupling of 1 with DDQ/Sc(OTf)3 yields 2 and 3,13 but substitution of the para fluoro groups 2 or 3 is inefficient. Adding the sugars to compound 1
proceeds in high yields, followed by oxidative coupling of 4b with PIFA efficiently yields 5a and 6a, which are readily deprotected to yield 5b and 6b.

S. Singh et al. / Tetrahedron Letters 55 (2014) 6311–6314
6313
Table 1
the electron withdrawing groups on 1 inhibit the oxidative cou-
pling and DDQ/Sc(OTf)3 reacts with the sugars, we employed the
hypervalent iodine(III) reagent, phenyliodine bis(trifluoroacetate)
(PIFA) for oxidative coupling.22 The oxidation of 1 bearing an open
meso position with different equivalents of PIFA results in the for-
mation of meso–meso linked diporphyrin 2 and the b–b, meso–
meso, b–b triply linked diporphyrins 3 (Scheme, ESI 2).23–27 The
metal ion chelated by the macrocycle improves the yields.
Porphyrin 1 was treated with 3.5 equiv of thioglucose and then
Zn(II) was inserted to efficiently yield 4b. The glycosylated triply
linked compound 5b is obtained in 40% yield after reacting with
2.5 equiv of PIFA, and 1.2 equiv yield predominately 6a. The struc-
tures of all porphyrinoids were confirmed by 1H, 19F and 13C NMR
spectroscopy, UV–visible, and MALDI-TOF spectra (ESI).
The UV–visible and emission spectra of the glycosylated deriv-
ative 5b and 6b are similar to those reported for 3 and 2, respec-
tively.13 Because the porphyrins are orthogonal in 6b, the
photophysical properties are similar to meso aryl porphyrins and
aggregates somewhat less than the fused derivative. However,
the UV–visible spectra 5b exhibit a broad Soret band at 413 nm
and Q-bands between 557 nm and 1068 nm; in different solvents
because the compound can partition into different cellular envi-
ronments (Table 1 and ESI). No apparent aggregation of 5b was
observed in DMSO, but in other solvents the compound aggregates
as indicated by shoulders on both blue (H-aggregate) and red (J-
aggregate) edges of the main absorption peaks.
The aggregation of 5b was confirmed by dynamic light scatter-
ing (DLS) measurements in these solvents (ESI Table 1). The size
and structural organization of nanoaggregates of porphyrinoids
depend on a variety of factors such as concentration, type of sol-
vent used, temperature, and nature of peripheral groups attached
to the chromophore.28 After shaking in phosphate buffered saline
(PBS), DLS shows two populations with diameters of 30 4 nm
and 284 15 nm, but after sonication for about 15 min, only
82 7 nm particles are observed. The size of the nanoaggregates
UV–visible spectral data for 5b
a
Solvent
UV–visible kmax (nm)
DMSO
429, 462, 569, 610, 636, 749, 876, 943, 1090
427, 462, 525, 565, 606, 667, 829, 966, 1079
428, 462, 524, 565, 606, 671, 848, 952, 1070
426, 461, 526, 565, 607, 671, 849, 950, 1071
421, 455, 514, 561, 602, 664, 837, 935, 1029, 1078
Toluene
Ethylacetate
PBS
Ethanol
a
1 cm path length. See ESI. In CHCl3 the lifetime is 3.3 ps, but no quantum yield is
reported (Ref. 13).
Our work on perfluorophenylporphyrins suggests that the triply
linked fused diporphyrin bearing six perfluorophenyl groups 5, and
singly bridged derivative 6 (Fig. 1) could serve as platforms to form
bioconjugates and biocompatible compounds thereby exploiting
the photophysical properties of 2 and 3 reported by Kim and co-
workers.8,13,17–20 Herein we report the synthesis, optical proper-
ties, cell uptake, and photodynamic induced necrosis activity of
hexa-glycosylated diporphyrin, 5b, and the fluorescence imaging
properties of 6b. Since the substitution of all six para fluoro groups
on 3 and isolation of this product proved difficult, we used the
trisperfluorophenyl porphyrin precursor (1, Fig. 1) as the platform
to append the thiosugars, and then formed the fused porphyrin by
a mild oxidative coupling reaction. The generality of the substitu-
tion chemistry of 1 affords a platform to append a diversity of con-
jugates, thereby broadening the applications of the two dimers.
Results and discussion
5,10,15-Tris(pentafluorophenyl)porphyrinatozinc(II) (1) was
synthesized using the procedure reported21 (Scheme ESI 1), and
is the platform molecule for the synthesis of singly and triply
linked diporphyrins with diverse biotargeting motifs. The oxidative
coupling reported by Osuka used DDQ/Sc(OTf)3 to form 3.13 Since
Figure 2. MDA-MB-231 cells were incubated with ca. 50 nM of the singly linked dimer 6b for 24 h, rinsed four times with PBS buffer to remove any unbound dye and fixed
with 4% paraformaldehyde solution in NaOH. Fluorescence images were captured using 540–580 nm band pass excitation and 600–650 nm band pass emission, magnification
20Â under identical instrumental conditions. (A) Just after preparation of the fixed cells, and (B) the same slide after 3 days. The contrast in each case was enhanced by 40% for
publication.

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S. Singh et al. / Tetrahedron Letters 55 (2014) 6311–6314
dimers add the palate of dyes for platforms for biomedical
applications.
Acknowledgments
Supported by the National Science Foundation – United States
(NSF) through CHE-0847997 to C.M.D. and DMR-0611539 to R.G.
Hunter College science infrastructure is supported by the NSF,
the National Institute on Minority Health and Health Disparities
– United States (8G12 MD007599) and the City University of
New York – United States.
Supplementary data
Supplementary data associated with this article can be found, in
the
online
version,
at
http://dx.doi.org/10.1016/j.tet-
let.2014.09.085. These data include MOL files and InChiKeys of
the most important compounds described in this article.
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and strength of intermolecular interactions significantly affect the
endocytosis of the photosensitizer by cancer cells, and the propen-
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of 6b are taken up by MDA-MB-231 human breast cancer cells and
slowly disaggregates inside the cells as indicated by the increased
fluorescence intensity (Fig. 2). Since 5b does not fluoresce, the dis-
aggregation is indicated by the PDT effects, wherein the light may
contribute to the disaggregation via internal conversion.
The singlet oxygen quantum yield (1O2) production (UD) for 5b
in DMSO was determined to be 0.78 0.03 in which the 1O2 phos-
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(4-sulfonatophenyl)porphyrin
dihydrochloride
(TSPP)
was
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5b and 6b were examined for PDT effect on the MDA-MB-231
human breast cancer cells (Fig. 3) where the percent of viable cells
is significantly reduced upon light exposure. Irradiation with white
light causes necrosis with an IC50 for 5b of ca. 13
lM and for 6b ca.
19 M measured just after photodynamic treatment, and 24 h
l
later, ca. 75% of the cell are necrotic for compound 5b and ca.
66% necrotic for compound 6b, indicating induction of apoptosis.14
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