1. BIOCOMPATIBILITY STUDIES Original Research
Nanosilver based anionic linear globular dendrimer with a special
significant antiretroviral activity
Mehdi Shafiee Ardestani1 • Alireza Salehi Fordoei2,3 • Asghar Abdoli2 •
Reza Ahangari Cohan4 • Golnaz Bahramali2 • Seyed Mehdi Sadat2 •
Seyed Davar Siadat5 • Hamid Moloudian6 • Nasser Nassiri Koopaei7 •
Azam Bolhasani2 • Pooneh Rahimi2 • Soheila Hekmat2 • Mehdi Davari2 •
Mohammad Reza Aghasadeghi2
Received: 10 January 2015 / Accepted: 30 March 2015
Ó Springer Science+Business Media New York 2015
Abstract HIV is commonly caused to a very complicated
disease which has not any recognized vaccine, so designing
and development of novel antiretroviral agents with
specific application of nanomedicine is a globally inter-
ested research subject worldwide. In the current study, a
novel structure of silver complexes with anionic linear
globular dendrimer was synthesized, characterized and
then assessed against HIV replication pathway in vitro as
well. The results showed a very good yield of synthesis (up
to 70 %) for the nano-complex as well as a very potent
significant (P 0.05) antiretroviral activity with non-sev-
ere toxic effects in comparison with the Nevirapine as
standard drug in positive control group. According to the
present data, silver anionic linear globular dendrimers
complex may have a promising future to inhibit replication
of HIV viruse in clinical practice.
Abbreviations
HIV Human immunodeficiency virus
RSV Respiratory syncytial virus
PEG-600 Polyethylene glycole 600 Da
DCC Dicyclohexyl carbodiimide
DLS Dynamic light scattering
DMSO Dimethyl sulfoxide
FTIR Fourier transforms spectroscopy
XTT 2,3-Bis(2-methoxy-4-nitro-5-sulfophenyl)-2H-
tetrazolium-5-carboxanilide
MPPI Mannosylated poly (propyleneimine)
dendrimer
1 Introduction
Globally, more than 40 million people suffer from the
human immunodeficiency virus (HIV) infection. The high
activity antiretroviral therapy (HAART) combining three
or more of antiretroviral (ARV) drugs administered for
lifelong renders acquired immunodeficiency syndrome
(AIDS) as a controlled chronic disease [1]. Limitations of
the current ARV include long lasting medications with
remarkable adverse effects, poor patient-medication ad-
herence, multi-drug resistance (MDR) due to mutations,
drug–drug interactions, poor drug pharmacokinetics, viral
load rise very quickly following medication discontinued
and affordability as well as establishing a latent infection
[2]. Unique properties of dendrimers as novel and exciting
& Mohammad Reza Aghasadeghi
mr_sadeqi@yahoo.com; mrasadeghi@pasteur.ac.ir
1
Department of Radiopharmacy, Faculty of Pharmacy, Tehran
University of Medical Sciences, Tehran, Iran
2
Department of Hepatitis and AIDS, Pasteur Institute of Iran,
13164 Tehran, Iran
3
Department of Biology, College of Basic Sciences, Tehran
Science and Research Branch, Islamic Azad University,
Tehran, Iran
4
Virology Research Group, Department of Rabies, Pasteur
Institute of Iran, Tehran, Iran
5
Department of Bacteriology, Pasteur Institute of Iran, Tehran,
Iran
6
Faculty of Pharmacy, Pharmaceutical Quality Assurance
Research Center, Tehran University of Medical Sciences,
Tehran, Iran
7
Department of Pharmaceutical Biotechnology, Faculty of
Pharmacy, Tehran University of Medical Sciences, Tehran,
Iran
123
J Mater Sci: Mater Med (2015) 26:179
DOI 10.1007/s10856-015-5510-7

2. polymers such as uniform size, water solubility, available
internal cavities, multivalency, well-defined molecular
weight, among others, make them attractive for drug de-
livery purposes since past decades [3–5]. These features are
caused by highly branched, spherical and nanometer di-
mension of dendrimers [4]. Furthermore, attempts have
been made to apply dendrimers to other areas such as
immunology and development of vaccines, antimicrobials
and antivirals [3]. Biocompatibility and release of drugs
from dendrimers are controversial issues [3]. Selection of
polymers has always been affected by issues such as
biodegradability, biocompatibility, bioadhesiveness while
respecting physical and biochemical properties [5]. Spheric
shape of dendrimers makes their structures much more
accurately controllable. They are indeed nano-sized mole-
cules with sizes between 10 and 100 nm [5].
Dendrimers due to aforementioned characteristics have
emerged as an important class of antiretroviral carriers [6].
Dendrimers as organic nano-platforms can act in
therapeutic and diagnostic applications [7]. To deliver a
therapeutic and non-toxic dose of drug to diseases, nano
and micro-carriers are efficient tools [8]. Increased effi-
ciency and decreased toxicity of anti-retroviral drug loaded
dendrimers were investigated in previous studies. Dutta
and Jain studied targeting and anti-HIV potential of lami-
vudine loaded mannosylated poly (propyleneimine)
(MPPI) dendrimer and concluded that the drug release
profile and cellular uptake of drug were improved sig-
nificantly. MPPI is a surface modified version of poly
(propyleneimine) (PPI) [9]. Passive targeting of drug
loaded nano-carriers is limited to tumors, the reticular en-
dothelial system and lymph nodes. Attractive features of
dendrimers like narrow molecular weight distribution,
small size and easy incorporation of targeting ligand ren-
ders them remarkable applicability in antiretroviral therapy
[10]. Dendrimers are also advantageous with respect to
their drug interactions [11]. Dendrimers could be widely
used in targeted and controlled release drug delivery sys-
tems; however, toxicity problems still exist and need some
modifications. Release profile of dendrimers is easily
modifiable and controllable [12]. Citric acid–polyethylene
glycol–citric acid (CPEGC) dendrimers were mentioned as
biocompatible compounds and investigated as drug deliv-
ery systems. Some small and hydrophobic molecules and
drugs were incorporated into the above dendrimer. These
dendrimers had good water solubility, biocompatibility and
low toxicity [13]. Synthesis of dendrimers has recently
been facilitated by the divergent ‘lego’ and the convergent
‘click’ approaches. Drugs could be physically loaded
within the dendrimer or chemically attached to the surface.
Ester-terminated dendrimers are more bioavailable than
amino-terminated ones. Increase of dendrimer sizes makes
them more soluble. Nonetheless, cytotoxic effects of
dendrimers increase with their generation number. Cyto-
toxicity could be reduced by dendrimer functionalization
(particularly with PEG). Dendrimers can inherently act
against tumors, bacteria and viruses [14].
AgNPs have been used for their antibiotic effects,
noteworthy that no cross-resistance with other antibiotics is
evident and efficacy against multi drug resistant strains [15,
16]. AgNPs show wide spectrum against different bacteria
and fungi [17–22] commercially used in health and im-
plantable biomaterial, general utilities and home appli-
ances. Moreover, antiviral effects of AgNPs have attracted
attention recently. AgNPs present remarkable antiviral
activity against HIV, hepatitis B, HSV, RSV, monkeypox
virus, tacaribe virus (TCRV), RNA viruses and adenovirus
type 3 [21, 23–26]. The supporting mechanism for AgNPs
antiretroviral effect is through glycoprotein 120 (gp120)
binding and prevention following the penetration and in-
terfering with intracellular stages. With the characteristics
mentioned, AgNPs are applied in a vaginal product to
prevent sexually transmitted diseases like HIV. However,
higher efficacy has been proposed with the concomitant
use of neutralizing antibodies [23]. Wide anti-infectious
properties of AgNPs as well as wound healing effects
makes silver suitable candidate for topical formulations
[20, 22, 27, 28].
Limitations of current antiretroviral drugs such as ex-
tensive first pass metabolism, astrointestinal degradation,
poor solubility, etc. lead investigation of novel drug de-
livery systems (NDDS) such as sustained and targeted drug
delivery systems. So, in the present study, a novel structure
of silver complexes with anionic linear globular dendrimer
was synthesized, characterized and then assessed against
HIV replication pathway in vitro.
2 Materials and methods
2.1 Materials
PEG 600 was purchased (from Merck,Germany). Dicy-
clohexylcarbodiimide, dimethylsulfoxide, citric acid and
silver were obtained (from Merck,Germany). Besides
pSPAX.2, pmzNL4-3 and pMD2.G plasmids were used for
transfection. In cellular studies, HEK293T cell line and
DMEM primary culture medium were used. HEK is an
abbreviation for human embryonic kidney cells. This cell
line was chosen for this research because it has superior
survival and proliferation compared to adult cells.
2.2 Synthesis of dendrimers G1 and G2
Synthesis of compound G1 of dendrimer was initiated
through the reaction of PEG 600 (as the core of dendrimer)
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123

3. and dicyclohexylcarbodiimide (DCC) in anhydrous DMSO
in the condition of darkness and vacuum. DCC activated
PEG by substitution with hydrogen atom of carboxylic acid
groups. Then, DCC was replaced with citric acid, and so
color of the tick solution changed to orange with stench.
Reaction was terminated with adding water and after fil-
tration; yellow-brown solution of dendrimer was obtained.
Primarily, synthesis of compound G1 was confirmed by
thin layer chromatography (TLC); stationary phase: silica
gel, mobile phase: 65 % CCl4/35 % MeOH. Then, the
product of G1 was separated and purified by dialysis.
Volume and concentration of dialysis buffer was 1600 fold
and half of the contents of dialysis bag, respectively. Pore
size of dialysis bag was 500–1000 Da. PEG was used as
limiting substrate of reaction and didn’t leave any residue.
Other substrates were excess. Mass of dendrimer was
transferred to several vials and lyophilized in yield 91 %
after freezing in liquid nitrogen. Procedures were severally
repeated to obtain the optimum procedure. To synthesize
dendrimer G2, DCC and citric acid were added to solution
of dendrimer G1 in DMSO and the solution became ticker
and thereafter dark cream color was appeared. In this stage,
compound G1, was rate limiting of reaction. As synthesis of
dendrimer G1, reaction was terminated with water addition,
solution was filtrated, dialyzed, and powder was lyophi-
lized. Structure of dendrimer G2 (in yield 73 %) was
confirmed by FT-IR. Size and electric charge of G1 and G2
dendrimers was determined by zetasizer (dynamic light
scattering, DLS).
Briefly, 2 mg of plasmid mixture was dissolved in
150 ll of deficient culture medium. Then, 40 ll of lipo-
fectin was added to the tube containing DNA and incubated
in room temperature for 10 min. Following that, 3 mL of
cell medium.
2.3 Preparation of Ag-dendrimer conjugates
Tendency of nano-silver particles to the oxygen atoms of
dendrimer leads to the reaction and incorporation of silver
into the dendrimers. In the isolated and light protected
condition, the mixture of dendrimer and silver was stirred
for 24 h. Yellow-brown color of mixture changed to gray.
Then, the mixture was filtered and brown solution of Ag-
dendrimer conjugated was separated from insoluble free
silver. Final product (liquid or lyophilized) was perma-
nently preserved from light and air, during the analyses.
Structure of product was assessed by FT-IR and compared
with G1 and G2 dendrimers. Size and charge of the final
product was determined and compared as well. Imaging of
Ag-dendrimer conjugated was made by AFM. Concentra-
tion of conjugated silver was determined by ICP-Mass
technique. Maximum incorporation of silver into the den-
drimer was calculated by ICP-Mass and divided by total
mass of G2 dendrimer and multiplied by 100; therefore
percentage of loading complex was obtained (52 %).
2.4 Cell culture and antiviral assessments
2.4.1 Cells, transfection and virus production
To produce HIV-1 SCR virions pseudotyped by VSVG,
Qiagen sample was used. First HEK293T cells were cul-
tured and maintained in Dulbecco’s modified Eagle’s cul-
ture medium (DMEM, Sigma, USA) supplemented with
FBS (15 %), penicillin (100 U/ml), streptomycin (100 lg/
ml), and glutamine (2 mmol/l); incubated in 5 % CO2,
37 °C inside a 6-well plate until they reach a total number
of approximately 1.2 9 106
. Transfection was carried out
using standard Lipofectamine 2000 according to Qiagen
protocol. Plasmids of pSPAX.2, pmzNL4-3 and pMD2.G
were simultaneously transfected into HEK293T cells to
produce proviruses. Next, cell culture plates were incu-
bated for minimum time of 24 h in 37 °C and 5 % CO2.
HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic
acid) (25 mM) was used to adjust the pH of the cell culture
medium in transfection stage. To prepare virus stock, cul-
ture supernatants of the transfected cells were harvested at
24, 48, and 72 h post transfection, and after filtration were
concentrated for 2 h in 60,0009g in 4 °C by ultracen-
trifugation [29]. The pelleted virions were solved in 1/20
volume of RPMI 1640 by mixing gently overnight at 4 °C
to prevent virus aggregation, quantified by P24 ELISA
assay kit (Biorad) and stored at -70 °C until use [30].
HEK cells were used to determine P24 titer of SCR HIV-1
virions pseudotyped by stomatitis virus surface VSVG
vesicular glycoprotein. A 60 9 103
count of HEK cells
were inoculated in each well of a 24 well plate containing
250 ll of culture medium and then 600 ng of P24 was
added to each well. Next, the cells were washed every 5 h
using 10 % FBS. Every time, the supernatants were col-
lected and replaced with new culture medium. Finally, P24
concentration in the supernatant was determined according
to the ELISA capture protocol (Biomerieux) developed by
the manufacturing company. All the components were
dissolved in dimethylsulfoxide (DMSO) with concentra-
tions ranging from 10 lM to 10 mM. Nevirapine (NVP)
was dissolved in DMSO with 50 lM concentration, ex-
tracted from commercial preparations was used as positive
control to inhibit viral growth and DMSO with final con-
centration of 1 % as the negative control. All the ex-
periments were triplicate. Toxicity of compounds was
separately estimated on the HEK cells. CC50 (cytotoxicity
capacity 50 %) was calculated. Dividing CC50 by IC50
provided TI (therapeutic index) and/or SI (selectivity in-
dex) (TI or SI = CC50/IC50). Antiviral effects of different
compounds were compared through TI or SI indices.
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123

4. 2.5 Evaluation of the new nano-drug effectiveness
on HIV replication
A HEK cells count of 60 9 103
in each well of a 24 well
plate containing 250 ll of culture medium was prepared.
SCR HIV virions and 600 ng P24 were added to each well.
The compounds were added in four concentrations (20, 40,
80 and 120 lM) to the wells as triplicate.
After 5 h, the cells were washed twice with DMEM/FBS
10 % [31]. Again, the compounds and fresh medium with
the same amounts were added. At 72 h after inoculation,
the supernatant was collected. The P24 level of the su-
pernatant was determined with ELISA capture (Biorad).
Related ELISA test was performed and finally, the con-
centration was calculated. To examine cellular prolif-
eration, XTT method (Roche) was used [30]. Medium of
phenol red was used to diminish test errors in proliferation
assay.
3 Results
3.1 Time relation of the SCR viral replication
Replication of the SCR HIV-1 virions psuedotyped by
VSVG on the HEK cells was determined after viral
inoculation. P24 secretion was started at the first day and
increased to a maximum level at 48 h after SCR virions
inoculation. Then, P24 level was gradually decreased from
the fourth day after inoculation. As the results denote,
maximum cumulative amount of P24 in the supernatant
was 72 h after inoculation. Therefore, P24 determination in
the supernatant was used to assess the viral replication at
72 h after inoculation (Fig. 1).
3.2 SCR HIV-1 replication regarding the primary
MOI
HEK cells were infected by different multiplicity of in-
fection (MOI) of pseudotype SCR virions. The supernatant
of the infected cells was assessed through P24 level 3 days
after inoculation. An inverse linear relationship between
MOI and P24 production makes this test a precise method
to investigate replication. Based on the assessments, the
adequate viral count for replication start is 600 ng P24
(Fig. 2).
3.3 Nevirapine efficacy as the control positive
Based on the results, suitable viral count for replication
(600 ng P24) and time (72 h) for P24 determination was
selected. To assess nevirapine function that is a NRTI drug,
SCR virions replication in the presence of its different
concentrations was measured. The results showed that in-
creasing nevirapine level decreased P24 production and
50,000 nM concentration inhibited P24 production. These
results revealed the correct function of the drug in the
primary test conditions and replication assessment based
on SCR HIV-1 suitably shows drug function in different
concentrations (Fig. 3).
3.4 Inhibitory effects of silver dendrimer
nanoconjugate
The ability of Iron chelating compounds to inhibit HIV
virus replication was investigated by assessing HIV repli-
cation in the presence of various compounds and calculated
based on the light absorption in P24 ELISA OD.
Nanoconjugated silver dendrimer effects in 20, 40, 80 and
120 lM concentrations on HEK cells was assessed. The
best antiviral effects were observed in 120 lM that inhib-
ited viral contamination up to 87 %. Generally speaking,
higher levels of drug produced higher level of viral
Fig. 1 The effects of nanosilver on the P24 levels at different time
exposures
Fig. 2 The effects of nanosilver on the different amounts of starter
virus
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5. inhibitory effects. This was also evident for control (silver
nitrate) (see more detailed data at Table 1, Fig. 4).
3.5 Cytotoxicity of nanoconjugated silver
dendrimers in different concentrations
Based on ODs from ELISA proliferation test, cytotoxicity
was assessed through XTT method after 72 h. Although,
the compounds showed potent antiviral effects but they had
cytotoxicity as well. Cell survival in concentrations of 40,
80 and 120 lM was 23, 36 and 48 %, respectively and was
50 % in 80 and 120 lM concentration of silver nitrate as
the control. DMSO was used as the positive control. In-
hibitory effect was low at 20 lM concentration while low
cytotoxicity was observed as well (see more detailed data
at Table 2, Fig. 5).
3.6 TI indexing
According to the calculated IC50 and CC50, TI index was
determined for different compounds. Comparing this index
for various concentrations of nanoconjugated silver den-
drimers provides a method to identify concentrations with
higher antiretroviral effects and lower cytotoxicity.
Therapeutic index (TI) in 20 lM concentration was 5.3, the
best among all concentrations. TI for 40, 80 and 120 lM
concentrations was 0.96, 0.54 and 0.26, respectively.
Dendrimeric seizes for G1 and G2 products were 70 and
90 nm, respectively; dendrimeric charges were -3.5 and
-11.8 millivolt/Ampere (mv A-1
), respectively. More
Fig. 3 The effects of
Nevirapine on the P24 at
different time exposures
Table 1 Depiction of some
details on HIV inhibitory effects
of nanosilver and the control
drug
P24 OD Mean P24 OD HIV inhibitory %
Uninfected cells 0.04, 0.03 0.035 –
Virus control (without drug) 0.8, 0.9, 0.7 0.8 0
Nevirapine (anti-retroviral drug) 0.02, 0.05 0.03 100
Ag-dendrimer (120 lM) 0.08, 0.2 0.14 87
Ag-dendrimer (80 lM) 0.21, 0.32 0.26 67
Ag-dendrimer (40 lM) 0.42, 0.34 0.38 52
Ag-dendrimer (20 lM) 0.48, 0.88 0.68 15
AgNO3 (control) (120 lM) 0.04, 0.1 0.07 96
AgNO3 (control) (80 lM) 0.32, 0.42 0.37 53
AgNO3 (control) (40 lM) 0.55, 0.4 0.47 41
AgNO3 (control) (20 lM) 0.62, 0.92 0.77 4
Fig. 4 Antiretroviral effects of AgNO3 and silver based dendrimer
nanoparticle
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6. citric acid moieties at the surface of compound G2 leads to
its larger size and more negative charge compared with
compound G1. Atomic force microscopy (AFM) imaging
of dendrimer G2 was performed and compared with G1.
The important absorption bonds in FTIR spectrum for G1
are: 3000–3500 cm-1
for hydroxyl group of citric acid
attached to PEG, 2800–2900 cm-1
for aliphatic carbon
chain of citric acid and PEG, and shifted peak of carbonyl
group of citric acid from 1700 to 1600 cm-1
that confirms
low concentration of citric acid in the structure of den-
drimer and conjugation of citric acid and PEG. The ab-
sorbed bonds in FTIR spectrum [OH group of citric acid
attached to PEG (broad, 3500–3000 cm-1
), C=O
(1600 cm-1
)] for G2 are: wide and strong peak from 3000
to 3500 cm-1
for hydroxyl group of citric acid, strength-
ened peak from 2800 to 2900 cm-1
for aliphatic carbon of
citric acid and PEG and a strong peak about 1700 cm-1
is
related to strengthened steric bond of citric acid addition
[OH group of citric acid (broad and strong,
3500–3000 cm-1
), steric bond of citric acid (1700 cm-1
)].
Last mentioned peak implies high concentration of citric
acid in the structure of compound G2 and conjugation of
citric acid and PEG. Considering significant increase in
aliphatic and hydroxyl peaks of compound G2 compared
with G1; thus it can be concluded that IR spectra approved
the synthesis of both dendrimer generations.
4 Discussion
HIV transcripts are RNA to DNA by RT and introduce
provirus into the host cell DNA by integrase. CD4?
T cells
are the major hosts and with proceeding decrease of their
reservoir, opportunistic infections and cancers lead death.
HIV/AIDS has imposed heavy economic and health costs
to world health. Because of loss of vaccine, antiviral drugs
have been the best confronting approach. Kim et al. ex-
amined oral toxicity of AgNPs for 3 months finding the
target organ to be the liver with biliary tract hyperplasia,
possible necrosis, fibrosis and pigmentation. Finally, the
LOAEL and NOAEL were determined to be 125 and
30 mg/kg, respectively, while AgNPs could accumulate in
tissues investigated [32]. AgNPs exert bactericidal activity
through concentrations that no remarkable acute human
toxicity would emerge [22, 33]. But toxicity of available
antiviral drugs and resistance of virus increased the im-
portance of development of new antiretroviral drugs [34,
35]. Inhibition of virus replication via interaction with
cellular pathways is the current strategy [36–38].
Combination of nanotechnology and antimicrobial prop-
erties of Cr, Ag, Ti, and Zn presents effective anti-HIV ap-
proaches. Some studies have shown antiviral activities of
nano-silver (against HIV-1, HBV, HSV, RSV, and monkey-
pox virus) [39]. Some other studies showed nano-silver par-
ticles inhibit HIV fusion to the host cell via interaction with gp
120 [23, 40]. Furthermore nano-silver particles can interact
with HIV through other ways such as inhibition of reverse
transcriptase, protein activity and inhibition of host cell cycle.
Dendrimers can trap different molecules within their
branches and protect them from environmental influences
and controllably release in targeted positions. Also den-
drimers increase solubility of drugs by trapping them.
Table 2 Illustration of some
details on cell viability effects
of nanosilver and the control
drug
ELISA OD (450 nm) Mean OD Cell viability %
Un-treated (1 ll DMSO) cells 0.65, 0.61, 0.55 0.6 100
Ag-dendrimer (120 lg) 0.14, 0.14 0.14 23
Ag-dendrimer (80 lg) 0.22, 0.22 0.22 36
Ag-dendrimer (40 lg) 0.31, 0.28 0.29 48
Ag-dendrimer (20 lg) 0.49, 0.51 0.5 83
AgNO3 (control) (120 lg) 0.31, 0.29 0.3 50
AgNO3 (control) (80 lg) 0.32, 0.28 0.3 50
AgNO3 (control) (40 lg) 0.43, 0.45 0.44 73
AgNO3 (control) (20 lg) 0.53, 0.43 0.48 80
Fig. 5 Effects of different concentrations of Ag-dendrimer and
AgNO3 exposures on cell viability percentage results of
179 Page 6 of 8 J Mater Sci: Mater Med (2015) 26:179
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7. Dendrimers can carry molecules on their surface via in-
teraction with functional groups; a property that enables
specific cell or tissue targeting.
In this study we investigated several concentrations of
Ag-dendrimer nano-conjugated compounds against repli-
cation of HIV and determined cytotoxicity of the new
compound in different concentrations. Researches showed
the Ag-dendrimer conjugated reduces expression of P24
antigen on the surface of monocytes and lymphocytes. It
seems virus inhibition is due to the reduction of cellular
proliferation. Inhibition of cellular proliferation can lead
inhibition of P24 production in lymphocytes. As a result, it
seems Ag-dendrimer conjugated don’t target virus directly
and indeed attack host cell of replicating virus. But some
other drugs such as nevirapine inhibit replication, directly
without any influence on cellular proliferation. HIV repli-
cation rate and survival was assessed after application of
several concentrations (20, 40, 80, and 120 lM) of new
compound. Inhibition of cellular proliferation cause to in-
hibit replication of virus but clinical researches have shown
nano-silver particles decrease P24 level with 50 % without
any effect on cellular proliferation. In a mathematical
model for anti CD4?
therapy, it has been showed that 25 %
decrease in CD4?
cell counts can reduce function of virus
and rate of mutations [41]. With this respect current
compound by killing proliferating CD4?
T cells and de-
pleting host cell from dATP, inhibit P24 production and
HIV-1 replication [42–45]. Our previous studies have
demonstrated that biocompatible range of Anionic linear-
globular dendrimer up to the concentration of 0.5 mg/ml
and less of the Anionic linear-globular dendrimer-cis-
platinum(II) conjugates would be required to obtain a
similar therapeutic effect against different cancer cell lines
when compared with cisplatin treatment alone [46, 47].
Decreasing dose of drug usage by dendrimers may result in
less drug resistance and drug side effects.
Results of this study showed that all concentrations of
Ag-dendrimer nano-conjugated could inhibit HIV effec-
tively. Inhibition percent of HIV for concentrations 120,
80, 40, and 20 lM is 78, 67, 52, and 15, respectively. First
three concentrations have highest inhibition rates but show
high level of cytotoxicity as well (77, 64, and 52 %, re-
spectively). Although inhibition rate of concentration
20 lM was only 15 %; but its cytotoxicity was 17 % and
this concentration of new compound is totally more ap-
propriate. AgNO3 was used as control and concentration of
120 lM showed better inhibition and lower toxicity (96
and 50 %, respectively) than conjugated compound but not
appropriate for clinical application. At the concentration of
80 lM, the new compound is better than AgNO3 (53 %
inhibition), but its cytotoxicity is high (50 %). In this
concentration both compounds had remarkable cyto-
toxicity, too. At the concentration of 40 lM,
nanoconjugated compound had a stronger inhibition and
lower cell survival profile (48 % in front of 73 %) than
AgNO3. In this concentration none of compounds couldn’t
be selected as an antiretroviral drug. At the concentration
of 20 lM, AgNO3 has a very low inhibition rate (4 %) and
a considerable cytotoxicity (20 %), while Ag-dendrimer at
concentration of 20 % had a stronger inhibition and lower
cytotoxicity profile.
5 Conclusion
Finally, we conclude that the concentration of 20 lM of
nanoconjugated compound has the best therapeutic profile
(stronger inhibition and lower toxicity). Silver saturated
dendrimers were used in this study and highly toxic com-
pounds were obtained. Lower silver loaded dendrimers or
other generations of dendrimer such as G1 can lead to more
applicable and different results. Also we propose that the
combination of concentration of 20 lM dendrimer with
other antiretroviral drugs to be investigated for better
therapeutic profile.
Acknowledgments We thank all members of the Hepatitis and
AIDS Department and Department of Radiopharmacy, Faculty of
Pharmacy, Tehran University of Medical Sciences of Pasteur Institute
of Iran and for their advice and assistance with the experiments. This
work was supported by Grant No. 519 from Pasteur Institute of Iran,
Tehran, Iran.
Conflict of interest The authors disclose no potential conflicts of
interest.
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