Immobilization of zidovudine derivatives on the SBA-15 mesoporous silica and evaluation of their cytotoxic activity

Article abstract of DOI:10.1371/journal.pone.0126251

Novel zidovudine derivatives, able to be covalently conjugated to silica surface, have been obtained and grafted to SBA-15 mesoporous silica. Cytotoxic activity of the hybrid organic-inorganic (zidovudine derivatives-silica) systems against HeLa and KB cell lines has been analyzed. Addition of folic acid had a positive influence on the cytotoxicity. Up to 69% of HeLa and 65% of KB tumor cells growth inhibition has been achieved at low silica concentration used (10 μg/mL).

Full text of DOI:10.1371/journal.pone.0126251

RESEARCH ARTICLE
Immobilization of Zidovudine Derivatives on
the SBA-15 Mesoporous Silica and Evaluation
of Their Cytotoxic Activity
1
1☯
2☯
2☯
Dawid Lewandowski *, Marta Lewandowska , Piotr Ruszkowski , Anita Pińska
,
1
☯
Grzegorz Schroeder
1
Faculty of Chemistry, Adam Mickiewicz University, Poznan, Poland, 2 Faculty of Pharmacy, Poznan
University of Medical Sciences, Poznan, Poland
☯
These authors contributed equally to this work.
dawid_le@amu.edu.pl
*
a11111
Abstract
Novel zidovudine derivatives, able to be covalently conjugated to silica surface, have been
obtained and grafted to SBA-15 mesoporous silica. Cytotoxic activity of the hybrid organic-
inorganic (zidovudine derivatives-silica) systems against HeLa and KB cell lines has been
analyzed. Addition of folic acid had a positive influence on the cytotoxicity. Up to 69% of
HeLa and 65% of KB tumor cells growth inhibition has been achieved at low silica concen-
tration used (10 μg/mL).
OPEN ACCESS
Citation: Lewandowski D, Lewandowska M,
Ruszkowski P, Pińska A, Schroeder G (2015)
Immobilization of Zidovudine Derivatives on the SBA-
1
5 Mesoporous Silica and Evaluation of Their
Cytotoxic Activity. PLoS ONE 10(5): e0126251.
doi:10.1371/journal.pone.0126251
Academic Editor: Heidar-Ali Tajmir-Riahi, University
of Quebect at Trois-Rivieres, CANADA
Introduction
Received: February 12, 2015
Accepted: March 31, 2015
Published: May 5, 2015
Mesoporous silicas discovered by Mobil researchers in the early 1990s gave rise to a family of
materials that have found numerous applications. From among many morphologically differ-
ent silicas, the two MCM-41 and SBA-15, have been the most often used. Both represent hexag-
onal order of mesopores but differ in pore diameters, which for MCM-41 is in the range of 2.0–
Copyright: © 2015 Lewandowski et al. This is an
open access article distributed under the terms of the
Creative Commons Attribution License, which permits
unrestricted use, distribution, and reproduction in any
medium, provided the original author and source are
credited.
6
.5 nm [1] and for SBA-15–4.0–30.0 nm [2, 3]. MCM-41 is usually used as a carrier for smaller
molecules that are packed inside its pores [4, 5], whilst SBA-15 is more suitable for larger ones
like proteins [3, 6]. Both can undergo useful surface functionalization, wherein for the SBA-15
a wider variety of molecules is applicable.
Application of mesoporous silicas in the preparation of controlled drug release systems is
well known, especially for transportation of anticancer and anti-inflammatory drugs. However,
a vast majority of these systems rely on the adsorption properties of anticancer drugs and gate-
like structures located on the pore entrances [7] or on surface modifications [8]. Only in very
few systems a covalent conjugation of the drug to the silica surface takes place [9].
Physically adsorbed anticancer drugs need to be transported, using mesoporous silica carri-
ers, to the vicinity of target tumor cells and protected from premature release by different sti-
muli-sensitive moieties. Covalently bound drugs require endocytosis of silica particles by the
tumor cells, which already has been reported [10]. The addition of covalently conjugated folic
acid enhances particles uptake [10, 11].
Data Availability Statement: All relevant data are
within the paper and its Supporting Information files.
Funding: Funding for this work came from the
institution Narodowe Centrum Nauki (PL), www.ncn.
gov.pl/, grant number: 2011/03/B/ST5/01573. The
funding was received by author GS. The funders had
no role in study design, data collection and analysis,
decision to publish, or preparation of the manuscript.
Competing Interests: The authors have declared
that no competing interests exist.
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Cytotoxic Activity of the Immobilized AZT Derivatives
Nucleoside analogues, especially 2’,3’-dideoxynucleosides, have found broad application as
antiviral [12] and anticancer therapeutics [13–16], designed to mimic natural nucleosides. One
of these molecules, which play an important role in anticancer therapeutics is 3’-azido-3’-
deoxythymidine (AZT, zidovudine).
AZT has been developed as an antitumor drug, but later it was found to reveal antiretroviral
activity and proved to be a proper drug for the treatment of acquired immunodeficiency syn-
drome (AIDS) [12]. There are also examples of application of AZT as an anticancer agent
in therapy of colon cancer, especially in combination with cisplatin, methotrexate and
5
-fluorouracil [17, 18].
The mechanism of antiviral or anticancer action of AZT involves its intracellular conversion
to its active, 5’-triphosphate form, which is controlled by the cellular enzymes called kinases.
The compound 5’-triphosphate is a competitive inhibitor of enzymes involved in the replica-
tion process (HIV reverse transcriptase or DNA polymerase) and after incorporation to DNA
strand it becomes a chain terminator of the nascent DNA strand due to the lack of 3’-hydroxyl
group [19, 20].
In 2001, Sharpless and his co-workers described the concept and criteria for the so-called
‘click’ reactions: versatility, high yield, modularity, readily available starting materials as well as
simple reaction conditions. In 2002, Meldal and Sharpless [21, 22] independently, established
that the Huisgen 1,3-dipolar cycloaddition reaction between organic azides and terminal al-
kynes can be efficiently catalyzed by copper(I) ions and can take place at room temperature. As
a result, the 1,4-regioisomers of 1,2,3-triazole are formed. ‘Click’ chemistry has been recognized
as a valuable synthetic tool in the field of nucleosides and nucleotides since azido and alkyne
derivatives are readily accessible [23]. 1,4-Disubstituted 1,2,3-triazole ring is a stable element of
the conjugate structure and it is considered as a mimic of Z-amide bond. Moreover, it can serve
as an additional pharmacophore [24].
To the best of our knowledge, there is no report on zidovudine (AZT) having been success-
fully grafted onto the silica surface. The aim of this study was to covalently conjugate zidovu-
dine to SBA-15 mesoporous silica surface and analyze the cytotoxic activity of hybrid organic-
inorganic systems obtained. All intended aims had been successfully achieved.
Materials and Methods
Materials
All reagents and solvents used in the study were purchased from Sigma-Aldrich and used with-
2
ꢀ
-1
out further purification. SBA-15 mesoporous silica (8–11 nm pore diameter, 600 m g surface
area and 1–2 μm particle size) was purchased from ACS Material.
Synthesis of zidovudine derivatives
One molar equivalent of (3-isocyanatopropyl)triethoxysilane in THF was placed in an ice bath
and an appropriate amount of propargyl compound (1.1 eq. for propargylamine and 5.0 eq. for
propargyl alcohol) in the presence of small amounts of triethylamine (0.6 eq. for propargyla-
mine and 0.25 eq. for propargyl alcohol, respectively) was added through a dropping funnel
and the mixtures were stirred for 3 h. The amount of propargyl alcohol was much higher due
to its weaker nucleophilic properties. While using lower propargyl alcohol:isocyanate ratios,
the products of side reactions were detected, especially those of isocyanate self—addition reac-
tion. The next step was to reflux the mixtures for 3 h, allow them to cool to the room tempera-
ture and stir overnight. Solvents were evaporated and residues were purified using column
chromatography (ethyl acetate:n-hexane 3:1) resulting in colorless, viscous liquid and pale yel-
low solid for ‘oxy’-AZT and ‘aza’-AZT intermediates, respectively (Fig 1). TLC detection was
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Cytotoxic Activity of the Immobilized AZT Derivatives
Fig 1. The reaction of synthesis of ‘aza’ and ‘oxy’ intermediates products.
doi:10.1371/journal.pone.0126251.g001
performed using ethyl acetate:n-hexane 1:2 as a mobile phase. In both cases the yield was about
0% and the compounds were characterized by electrospray mass spectrometry (Micromass
ZQ spectrometer, Waters), melting point measurement (Mel-Temp 1002D), infrared spectros-
7
1
13
copy (FT-IR Bruker IFS 66v/S) and H and C NMR spectroscopy (VARIAN Mercury 300).
In the second step, both intermediates were conjugated with AZT by the1,3-dipolar Huisgen
cycloaddition in anhydrous conditions using copper(I) acetate as a catalyst and acetonitrile as a
solvent. Exact conditions in both cases were adjusted to achieve the best purity and yield. After
the synthesis, both products were purified using column chromatography (methanol:chloro-
+
form gradient), which gave pale green (Cu remnants) and pale orange solids for ‘oxy’-AZT
and ‘aza’-AZT (Fig 2). TLC detection was performed using methanol:chloroform 1:10 as a mo-
bile phase. The compounds were characterized by electrospray mass spectrometry (Micromass
ZQ spectrometer, Waters), melting point measurement (Mel-Temp 1002D), infrared spectros-
1
13
copy (FT-IR Bruker IFS 66v/S) and H and C NMR spectroscopy (VARIAN Mercury 300).
Immobilization of zidovudine derivatives on SBA-15 mesoporous silica
surface
Both derivatives were immobilized on the surface by themselves or together with folic acid. To
anchor ‘aza’-AZT and ‘oxy’-AZT on the surface, 70 mg of each of the compounds were
Fig 2. Synthesis reaction of the ‘aza’- and ‘oxy’-AZT derivatives.
doi:10.1371/journal.pone.0126251.g002
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Cytotoxic Activity of the Immobilized AZT Derivatives
Fig 3. The immobilization of zidovudine derivatives on SBA-15 mesoporous silica surface.
doi:10.1371/journal.pone.0126251.g003
dissolved in 10 ml of DMF in separate flasks and 100 mg of SBA-15 silica was added to each
flask. Suspensions were stirred at 80°C for 1 h and then allowed to cool to the room tempera-
ture and stirred overnight. Modified silica was filtered off, washed thoroughly with DMF and
dried. SBA-15+’aza’-AZT and SBA-15+’oxy’-AZT were obtained as white solids (Fig 3). The
accurate amounts of both derivatives anchored to the surface were calculated using elemental
analysis (elemental analyzer EL III, Vario). The presence of AZT derivatives on the silica sur-
face was additionally confirmed by the IR spectroscopy for 1.0 mg of the silica mixed with
2
00 mg of KBr and formed into a pellet.
In the second approach 300 mg of SBA-15 silica was suspended in 10 ml of toluene and
0 mg of (3-aminopropyl)trimethoxysilane (APTMS) was added. The mixture was refluxed for
h and then allowed to cool to the room temperature and stirred overnight. After that the silica
3
3
was filtered off, washed with toluene and dried. SBA-15+APTMS as a white solid was obtained.
In the next step 50 mg of folic acid (FA) was dissolved in 30 ml of DMF at slightly elevated tem-
perature (40–45°C), an excess of diisopropylcarbodiimide (DIC) (30 mg) was added and, after
1
5 min, SBA-15+APTMS was suspended. The mixture was stirred for 24 h and then filtered
off, washed thoroughly with DMF and dried. SBA-15+APTMS+FA as a pale yellow solid was
obtained. In the final step 70 mg of both derivatives, ‘aza’-AZT and ‘oxy’-AZT, were dissolved
in 10 ml of chloroform in separate flasks and 150 mg of SBA-15+APTMS+FA was added to
each of them. Mixtures were refluxed for 20 min and then allowed to cool and stirred over-
night. Both solids were filtered off, washed with chloroform and dried, yielding SBA-15
+
APTMS+FA+’aza’-AZT and SBA-15+APTMS+FA+’oxy’-AZT (Fig 4). Both products were
analyzed using elemental analysis and IR spectroscopy.
The cytotoxic activity of hybrid materials
Human cancer cells HeLa (cervical cancer cell line) and KB (carcinoma nasopharynx) were cul-
tured in RPMI 1640 medium. Each medium was supplemented with 10% fetal bovine serum,
1
% L-glutamine and 1% penicillin/streptomycin solution. The cell lines were kept in the incu-
4
bator at 37°C. The optimal plating density of cell lines was determined to be 5 x 10 . All the cell
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Cytotoxic Activity of the Immobilized AZT Derivatives
Fig 4. Immobilization of folic acid and both zidovudine derivatives on SBA-15 mesoporous silica
surface.
doi:10.1371/journal.pone.0126251.g004
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Cytotoxic Activity of the Immobilized AZT Derivatives
lines were obtained from The European Collection of Cell Cultures (ECACC) supplied by
Sigma-Aldrich (catalogue numbers: HeLa cell line-93021013, KB cell line-94050408).
The protein-staining sulforhodamine B (SRB, Sigma-Aldrich) microculture colorimetric
assay, developed by the National Cancer Institute (USA) for in vitro antitumor screening was
used in this study, to estimate the cell number by providing a sensitive index of total cellular
protein content, which is in a linear relationship to the cell density [25]. The monolayer cell
4
culture was trypsinized and the cell count was adjusted to 5 x 10 cells. In each well of the 96
well microtiter plate, 0.1mL of the diluted cell suspension (approximately 10,000 cells) was
placed. After 24 hours, when a partial monolayer was formed, the supernatant was washed out
and 100 μL of six different silica suspension concentrations were added to the cells in microtiter
plates. The tested silicas were suspended in DMSO (20 μM) and the content of DMSO did not
exceed 0.1% as this concentration was found to be nontoxic to the cell lines. The cells were ex-
posed to silicas for 72 hours. After that, 25 μL of 50% trichloroacetic acid were added to the
wells and the plates were incubated for 1 hour at 4°C. The plates were then washed out with
distilled water to remove traces of medium and next dried by air. The air-dried plates were
stained with 100 μL SRB and kept for 30 minutes at room temperature. The unbound dye was
removed by rapidly washing with 1% acetic acid and then air dried overnight. The optical den-
sity was read at 490 nm. All cytotoxicity experiments were performed three times. Cell survival
was measured as the percentage absorbance compared to the control (non-treated cells). Zido-
vudine (Sigma-Aldrich) was used as the internal standard.
Results and Discussion
Characterization of synthesized compounds
Both propargyl-containing intermediates were obtained in high purity (full spectra available in
the supplementary material):
1
‘
aza’-AZT intermediate—melting point: 35–38°C, H NMR (300 MHz, CD CN) δ: 0.62–
3
0
3
.67 (2H, m), 1.21–1.25 (9H, t, J = 7.0 Hz), 1.58–1.66 (2H, m), 2.22 (1H, t, J = 2.5 Hz), 3.15–
.21 (2H, dt, J = 6.9 Hz, J = 5.9 Hz), 3.80–3.85 (6H, q, J = 7.0 Hz), 3.97–4.00 (2H, dd, J = 2.5
13
Hz, J = 5.5 Hz), 5.08–5.15 (2H, m), C NMR (75 MHz, CDCl ) δ: 7.5, 18.2, 23.5, 30.0, 42.8,
5
3
-
1
-1
8.4, 70.9, 80.8, 157.9, FT-IR: 3330 cm (N-H ν ), 3250 cm (ꢁC-H ν ), 2975, 2925 and 2885
st
-1 -1 -1 -1
st
cm (aliphatic C-H ν ), 2115 cm (CꢁC ν ), 1625 cm (urea C = O ν ), 1570 cm (N-H δ ),
st
w
st
st
-
1
+
+
+
1
080 cm (Si-O ν ), ESI MS m/z [M-C H O] 257.3, [M+Na] 325.3, [M+K] 341.3.
vs 2 5
1
13
‘
oxy’-AZT intermediate—melting point: below -25°C, H NMR and C NMR data can be
-
1
-1
found in literature [26], FT-IR: 3340 cm (N-H ν ), 3315 cm (ꢁC-H ν ), 2975, 2930 and
st
st
-
1
-1
-1
-1
2
890 cm (aliphatic C-H ν ), 2130 cm (CꢁC ν ), 1715 cm (carbamate C = O ν ), 1535 cm
st
-1 -1 + +
w
st
(
N-H δ ), 1235 cm (C-O ν ), 1080 cm (Si-O ν ), ESI MS m/z [M-C H O] 258.2, [M+Na]
st st vs 2 5
+
3
26.3, [M+K] 342.3.
aza’-AZT final product—melting point: 85–88°C, H NMR (300 MHz, CD CN) δ: 0.53–
1
‘
3
0
2
.58 (2H, m), 1.14–1.19 (9H, t, J = 7.0 Hz), 1.44–1.53 (2H, m), 1.85–1.86 (3H, d, J = 1.2 Hz),
.60–2.68 (1H, m), 2.76–2,83 (1H, m), 3.02–3.08 (2H, dt, J = 6.9 Hz, J = 6.0 Hz), 3.47–3.51 (1H,
t), 3.66–3.72 (1H, m), 3.74–3.84 (7H, m), 4.27–4.31 (1H, dt, J = 3.1 Hz, J = 5.4 Hz), 4.31–4.33
2H, d, J = 5.8 Hz), 5.11–5.15 (1H, t, J = 5.7 Hz), 5.26–5.31 (1H, dt, J = 5.4 Hz, J = 8.5 Hz),
.36–5.41 (1H, t, J = 5.7 Hz), 6.38–6.42 (1H, t, J = 6.6 Hz), 7.65–7.66 (1H, q, J = 1.2 Hz), 7.75
(
5
1
3
(
1H, s), 9.10 (1H, s), C NMR (75 MHz, CDCl ) δ: 7.6, 12.4, 18.2, 23.6, 35.5, 37.9, 43.0, 58.4,
3
-
1
5
3
9.4, 61.3, 85.2, 86.4, 110.9, 122.7, 137.1, 146.5, 150.8, 158.9, 164.6, FT-IR: 3320 cm (N-H ν ),
m
-
1
-1
-1
065 cm (= C-H ν ), 2975, 2925 and 2890 cm (aliphatic C-H ν ), 1700 cm (pyrimidine
m
st
-
1
-1
C = O ν and urea C = O ν as a distortion of this signal), 1560 cm (N-H δ ), 1275 cm
st
st
st
+
-
1
+
(
sugar C-O ν ), 1080 cm (Si-O ν ), ESI MS m/z [M+Na] 592.4, [2M+Na] 1161.7.
m vs
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Cytotoxic Activity of the Immobilized AZT Derivatives
1
‘
oxy’-AZT final product—melting point: 56–59°C, H NMR (300 MHz, CD CN) δ: 0.53–
3
0
2
.59 (2H, m), 1.14–1.19 (9H, t, J = 7.0 Hz), 1.47–1.56 (2H, m), 1.85–1.86 (3H, d, J = 1.2 Hz),
.61–2.69 (1H, m), 2.77–2.84 (1H, m), 3.03–3.09 (2H, q, J = 6.8 Hz), 3.31 (1H, s), 3.68–3.73
(
5
1H, dd, J = 3.1 Hz, J = 12.3 Hz), 3.74–3.84 (7H, m), 4.29–4.33 (1H, dt, J = 3.1 Hz, J = 5.4 Hz),
.10 (2H, s), 5.29–5.34 (1H, dt, J = 5.3, J = 8.5 Hz), 5.61–5.67 (1H, t, broad signal), 6.38–6.43
13
(
1H, t, J = 6.6 Hz), 7.64–7.65 (1H, q, J = 1.2 Hz), 7.88 (1H, s), 9.01 (1H, s), C NMR (75 MHz,
CDCl ) δ: 7.5, 12.4, 18.2, 23.1, 37.6, 43.4, 57.6, 58.4, 59.1, 61.2, 85.2, 87.6, 111.0, 124.2, 137.5,
3
-
1
-1
1
2
(
43.8, 150.5, 156.2, 164.2, FT-IR: 3330 cm (N-H ν ), 3065 cm (= C-H ν ), 2975, 2930 and
m m
890 cm (aliphatic C-H ν ), 2105 cm (very weak—probably unreacted AZT), 1700 cm
pyrimidine C = O ν and carbamate C = O ν ), 1535 cm (N-H δ ), 1275 cm (sugar C-O
st st st
ν ), 1080 cm (Si-O ν ), ESI MS m/z [M+Na] 593.3, [2M+Na] 1163.5.
-
1
-1
-1
st
-
1
-1
-
1
+
+
m
vs
Characterization of modified silica obtained
Mean amounts of nitrogen and carbon in SBA-15+’aza’-AZT equal to, respectively, 0.471%
and 1.379% permitted calculation of the total amount of ‘aza’-AZT on the silica surface. As the
content of carbon is slightly dependent on the number of Si-O-Si links between ‘aza’-AZT and
silica surface, the nitrogen percentage is more reliable and gives 2.735±0.185% mass percentage
of ‘aza’-AZT, that is 48.1±3.3 μmol of ‘aza’-AZT per 1g of modified silica. Similar calculations
for SBA-15+‘oxy’-AZT, with 0.374% of nitrogen and 1.247% of carbon content, lead to 2.538
±0.058% mass percentage or 44.5±1.0 μmol of ‘oxy’-AZT per 1 g of modified silica. The IR
Fig 5. IR spectra of all hybrid systems obtained and pure SBA-15 mesoporous silica for comparison.
doi:10.1371/journal.pone.0126251.g005
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Cytotoxic Activity of the Immobilized AZT Derivatives
spectra (Fig 5) indicate the presence of both zidovudine derivatives, which can be proved by
-
1
the presence of C = O amide stretching signal at 1659 cm , other signals are invisible, because
of the low derivative content. The shift of the signal assigned to the C = O bond can be con-
nected with the immobilisation process. Moreover, this signal in the spectra of zidovudine, ac-
-
1
cording to the literature [27–31], can be found between 1700 and 1650 cm .
IR spectra of the systems containing folic acid are more complicated. However, the main
-
1
signals can still be assigned, the signal at1654 cm is assigned to the stretching of C = O amide
-
1
from zidovudine, the signals at 1698 and 1609 cm come from C = O carboxyl and C = O
amide groups of folic acid respectively. The shift of the peak assigned to the stretching of O-H
-
1
-1
from 3425–3420 cm for the silicas without folic acid to 3370–3360 cm for both systems with
folic acid is thus also related to the presence of folic acid. Elemental analysis results for both
modified silicas are 3.428% of nitrogen and 9.019% of carbon for SBA-15+APTMS+FA+’oxy’-
AZT and 3.703% of nitrogen and 9.237% of carbon for SBA-15+APTMS+FA+’aza’-AZT. They
are difficult to interpret because of the number of steps in the synthesis. Nevertheless, the mass
increase correlates well with the elemental analysis results for simple SBA-15+‘aza’-AZT and
SBA-15+’oxy’-AZT systems and following this line of reasoning it can be calculated that in
SBA-15+APTMS+FA+’aza’-AZT and SBA-15+APTMS+FA+’oxy’-AZT systems the amounts
of zidovudine derivatives are slightly above 100 μmol per 1 g of modified silica, which is about
twice as much as for simple SBA-15+’aza’-AZT and SBA-15+’oxy’-AZT systems. Small
amounts of APTMS used in the first step of the synthesis left enough free space on the surface
Fig 6. Cytotoxic activity against HeLa cell line, calculated for all samples tested.
doi:10.1371/journal.pone.0126251.g006
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Cytotoxic Activity of the Immobilized AZT Derivatives
Fig 7. Cytotoxic activity against KB cell line, calculated for all samples tested.
doi:10.1371/journal.pone.0126251.g007
to anchor both derivatives, moreover the use of another solvent (of a lower dielectric constant
to facilitate its penetration into the silica [32]) brought a positive effect on their total amounts.
The cytotoxic activity
All analyzed solids showed concentration-dependent cytotoxic activity against HeLa and KB
cell lines (Fig 6 and Fig 7). Introduction of amino groups on the silica surface greatly
increased its activity, which is in agreement with the results observed for pure, organic com-
pounds [33–35].
Covering the surface with folic acid had ambivalent influence. On the one hand, modifica-
tion of the amino groups with folic acid causes a decrease in the activity of SBA-15+APTMS
+
FA, because the access to primary amino groups is blocked with folic acid molecules. On the
other hand, the addition of folic acid molecules to both systems containing zidovudine deriva-
tives, significantly increases their activity, promoting the interactions between silica particles
and cell membrane, as KB and HeLa cell lines are known to be FR-positive [36]. Higher cell
growth inhibition observed for SBA-15+APTMS+FA+’oxy’-AZT in comparison with the result
for SBA-15+APTMS+FA+’aza’-AZT may be due to the fact that carbamate group in the ‘oxy’
derivative is more susceptible to hydrolysis than the urea-like one in the ‘aza’ derivative, that
means it can release more zidovudine molecules from the surface. Cytotoxic activities obtained,
especially that of SBA-15+APTMS+FA+’oxy’-AZT, are comparable to those of pure zidovu-
dine(IC50 = 3.12 μg/mL for KB and 2.28 μg/mL for HeLa cell line), but at much lower zidovu-
dine concentration. High deviations of the results obtained for some of the samples are a result
of sedimentation in silica suspensions.
Conclusions
Two novel zidovudine derivatives have been obtained and characterized. Both of them have
been easily introduced onto the SBA-15 mesoporous silica surface using the grafting technique.
Cytotoxic activity against HeLa and KB cell lines of all solids obtained has been evaluated. The
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Cytotoxic Activity of the Immobilized AZT Derivatives
addition of folic acid on the silica surface enhances the activity of the derivatives studied due to
the interaction with folate receptors. The values of cell growth inhibition obtained for SBA-15
+
APTMS+FA+’oxy’-AZT are relatively high and equal to 69% and 65% for HeLa and KB
tumor cells. The results of this study can be a basis for further attempts at covalent conjugation
of cytotoxic compounds to the silica surface.
Supporting Information
S1 Spectrum. Full ESI MS spectra of ‘aza’- and ‘oxy’-AZT intermediates.
(
TIF)
S2 Spectrum. Full FT-IR spectra of ‘aza’- and ‘oxy’-AZT intermediates.
TIF)
S3 Spectrum. H NMR spectrum of ‘aza’-AZT intermediate.
TIF)
S4 Spectrum. C NMR spectrum of ‘aza’-AZT intermediate.
TIF)
S5 Spectrum. Full ESI MS spectra of ‘aza’- and ‘oxy’-AZT final products.
TIF)
S6 Spectrum. Full FT-IR spectra of ‘aza’- and ‘oxy’-AZT final products.
TIF)
S7 Spectrum. H NMR spectrum of ‘aza’-AZT final product.
TIF)
S8 Spectrum. C NMR spectrum of ‘aza’-AZT final product.
TIF)
S9 Spectrum. H NMR spectrum of ‘oxy’-AZT final product.
TIF)
S10 Spectrum. C NMR spectrum of ‘oxy’-AZT final product.
TIF)
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Author Contributions
Conceived and designed the experiments: DL GS. Performed the experiments: DL ML PR AP.
Analyzed the data: DL GS. Contributed reagents/materials/analysis tools: PR GS. Wrote the
paper: DL.
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