Pure Drug and Polymer Based Nanotechnologies For The Improved Solubility, Stability

ARTICLE IN PRESS
ADR-11952; No of Pages 12
Advanced Drug Delivery Reviews xxx (2009) xxxxxx
Contents lists available at ScienceDirect
Advanced Drug Delivery Reviews

j o u r n a l h o m e p a g e : w w w. e l s ev i e r. c o m / l o c a t e / a d d r
Puneet Sharma, Sanjay Garg
School of Pharmacy, The University of Auckland, Auckland, New Zealand
OF
Pure drug and polymer based nanotechnologies for the improved solubility, stability,
bioavailability and targeting of anti-HIV drugs
a r t i c l e
i n f o
a b s t r a c t
RO
Article history:
Received 14 June 2009
Accepted 14 September 2009
Available online xxxx
The impact of human immunodeciency virus (HIV) infection has been devastating with nearly 7400 new
infections every day. Although, the advent of highly active antiretroviral therapy (HAART) has made a
tremendous contribution in reducing the morbidity and mortality in developed countries, the situation in
developing countries is still grim with millions of people being infected by this disease. The new
advancements in the eld of nanotechnology based drug delivery systems hold promise to improve the
situation. These nanoscale systems have been successfully employed in other diseases such as cancer, and
therefore, we now have a better understanding of the practicalities and technicalities associated with their
clinical development. Nanotechnology based approaches offer some unique opportunities specically for the
improvement of water solubility, stability, bioavailability and targeting of antiretroviral drugs. This review
presents discussion on the contribution of pure drug and polymer based nanotechnologies for the delivery
anti-HIV drugs.
2009 Elsevier B.V. All rights reserved.
Keywords:
HIV
Polymer
Nanoparticle
DP
6
7
8
9
10
12
11
13
Q2
14
15
16
17
32
Introduction . . . . . . . . . .
Pure drug nanoparticles . . . .
Polymer based nanotechnologies
3.1.
Polymeric micelles. . . .
3.2.
Polymeric nanoparticles .
3.3.
Dendrimers . . . . . . .
4.
Conclusion and future directions
5.
Uncited reference . . . . . . .
References . . . . . . . . . . . . .
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
EC
1.
2.
3.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
RR
36
37
38
39
40
41
42
Q7 43
44
Contents
45
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
CO
35
34
1. Introduction
47
Human immunodeciency virus (HIV) related acquired immune

deciency syndrome (AIDS) has claimed over 25 million lives since its
discovery in 1981. Based on the profound knowledge gained about the
HIV replication cycle, several drug targets have been identied over
the years and effective treatment options are currently available [1].
The current clinical therapy, known as highly active antiretroviral
50
51
52
UN
46
48
49
31
30
TE
33
This review is part of the Advanced Drug Delivery Reviews theme issue on
Nanotechnology Solutions for Infectious Diseases in Developing Nations.
Corresponding author. School of Pharmacy, The University of Auckland, Private Bag
92019, Auckland, New Zealand. Tel.: +64 9 373 7599x82836; fax: +64 9 367 7192.
E-mail address: s.garg@auckland.ac.nz (S. Garg).
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
18
19
20
21
22
23
24
25
26
27
28
29
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
.
0
0
0
0
0
0
0
0
0
treatment or HAART, is considered as one of the most signicant

advances in the eld of HIV therapy [2]. Since mid 1990s, HAART has
made a remarkable contribution towards reducing the mortality in
patients.
Recommended HAART regimens include at least three actives [3].
Since majority of antiretrovirals are administered orally, their
adequate systemic absorption from gastrointestinal (GI) tract is a
prerequisite for successful therapy. Amidon et al. [4] suggested a
Biopharmaceutic Classication System (BCS) that identies the
solubility and permeability of drug substances as markers for their
oral bioavailability. Following BCS guidelines, a drug substance is
considered highly soluble when its highest dose strength solubilizes
in 250 ml or less of aqueous media over a pH range of 1.07.5 at 37 C
[5]. Likewise, a drug substance is considered highly permeable when
0169-409X/$ see front matter 2009 Elsevier B.V. All rights reserved.
doi:10.1016/j.addr.2009.11.019
Please cite this article as: P. Sharma, S. Garg, Pure drug and polymer based nanotechnologies for the improved solubility, stability,
bioavailability and targeting of anti-HIV drugs, Adv. Drug Deliv. Rev. (2009), doi:10.1016/j.addr.2009.11.019
53
54
55
56
57
58
59
60
61
62
63
64
65
66
ARTICLE IN PRESS
78
79
80
81
82
83
84
85
86
87
88
89
90
91
92
93
94
95
96
97
98
99
100
101
102
103
104
105
106
107
108
109
110
111
t1:1
t1:2
t1:3
t1:4
t1:5
t1:6
t1:7
t1:8
t1:9
t1:10
t1:11
t1:12
t1:13
t1:14
t1:15
t1:16
t1:17
t1:18
Q1 t1:19
2. Pure drug nanoparticles
132
In a pure drug nanoparticle formulation, submicron size particles

(mean particle size b1 m and 99th percentile particle size of b5 m)
of drugs are stabilized in aqueous medium with GRAS (generally
recognized as safe) stabilizers. Such formulation can be used for drugs
with properties such as insolubility in both water and oil, high melting
point, high log P and high dose [18]. Following NoyesWhitney
equation, progressive size reduction of the drug particles leads to an
increase in the surface area resulting in an increased dissolution rate.
Additionally, particle size reduction results in the decrease of the
diffusion layer thickness surrounding the particles [19] and an
increased concentration gradient between the surface of the particle
and bulk solution, which facilitates particle dissolution by increasing
dissolution velocity. Therefore, nanosizing is a suitable approach for
increasing bioavailability of those drugs where dissolution is the rate
limiting step in systemic absorption.
Van Eerdenbrugh et al. investigated the dissolution and in vitro
absorption of a poorly water soluble non-nucleoside reverse transcriptase inhibitor (NNRTI), loviride (water solubility 0.1 mg/l), after
nanonization [20]. In addition to poor water solubility, loviride has a
high melting point (225 C) and poor solubility in oils. Such
physicochemical properties make loviride a brick-dust molecule
with low oral bioavailability. Media milling led to mean particle sizes
133
OF
76
77
RO
74
75
112
113
TE
73
affects the quality of patient's life but also signicantly adds to the
economic burden of the health care system.
In the context of oral drug delivery, the important characteristics
of a molecule that needs to be considered for positive anti-HIV effects
are (i) solubility and ionization, (ii) lipophilicity and permeability,
(iii) stability in biological uids, (iv) gastrointestinal metabolism and
(v) viral reservoir targeting. When these properties are unfavorable
for drug development, alternative processing and formulation specic
approaches can be employed to attain maximum therapeutic gains.
The nanometer size and high surface area to volume ratio which affect
the pharmacokinetics and biodistribution of the associated drug
molecule are main features of nanotechnology based drug delivery
systems. The nanotechnology based approaches discussed in this
review for the delivery of anti-HIV drugs include pure drug
nanoparticles, polymeric micelles, dendrimers and polymeric nanoparticles. Several reviews are available on various pharmaceutical
aspects of these nanoparticulate systems such as preparation
methods, physicochemical properties, toxicity and others ([1117]).
This review presents information pertaining to their applicability
specically for anti-HIV drugs.
RR
EC
71
72
114
115
116
117
118
119
120
121
122
123
124
125
126
127
128
129
130
131
134
135
136
137
138
139
140
141
142
143
144
145
146
147
148
149
150
151
152
153
154
Table 1
Antiretroviral drugs in the WHO essential core drug list classied according to BCS (adapted from [7]).
Drug
CO
69
70
the extent of the intestinal absorption (parent drug plus metabolites)

in humans is determined to be 90% of an administered dose based
on a mass balance determination or in comparison to an intravenous
(i.v.) reference dose [5]. However, there are other factors such as
effect of efux and absorptive transporters, intestinal metabolizing
enzymes (CYP34A and UDP-glucuronosyltransferases (UGTs)) and
food that have to be taken into consideration for the appropriate
estimation of the bioavailability [6]. Considering properties such as
highest dose strength, dose number, solubility and log P, Kasim et al.
[7] have classied twelve antiretrovirals (12th edition of World
Health Organization Essential Core Drug List) in different BCS classes
(Table 1). Out of these twelve antiretrovirals, nine have either
solubility (BCS Class 2) or permeability (BCS Class 3) related issues.
Consequently, oral absorption and, in turn, bioavailability is variable
(Table 2) and mainly dependent on (i) stability in stomach acid
conditions, (ii) aqueous solubility and dissolution from dosage form
and (iii) permeability through the intestinal membrane. Most of the
protease inhibitors (Pis) have high molecular weight (N500 Da), pH
dependent solubility (high solubility at acidic pH) and high log P (high
lipophilicity) (Table 3), and all these properties adversely affect the
oral bioavailability [8]. In addition, they are substrates of Pglycoprotein efux transporters. Similarly, although the majority of
nucleoside reverse transcriptase inhibitors (NRTIs) show good
systemic absorption (N70% at clinically relevant doses), didanosine
(ddI) and zidovudine (AZT) show variable bioavailability. ddI is
unstable at acidic pH and is subjected to rst-pass metabolism; which
may be responsible for its lower bioavailability. AZT also undergoes
rst-pass metabolism and additionally shows dose dependent effect
on bioavailability.
HAART is a lifelong necessity and any non-compliance leads to a
rapid increase in the viral load [2]. The reason for this relapse is related
to the poor targeting ability of the antiretroviral agent to the latent
sites of infection [9]. Another major limitation of the current HIV
treatment is linked to the short residence time and resulting low
concentration of antiretroviral drugs at certain inaccessible viral
reservoir sites such as lymphatic system, macrophages, lymphocyte,
central nervous system (CNS) and lungs [10]. Thus, administration of
higher doses is required for prolonged duration to eradicate the virus.
This prolonged therapy is often associated with the emergence of
resistant HIV strains. In addition, there are major issues of adverse
drug reaction and drugdrug interaction associated with antiretroviral therapy. Although newer regimens in HAART have signicantly
lower toxicity, modulation of lipid and glucose metabolism is still a
major issue [2]. After chronic treatment even moderate toxicity may
lead to serious complications. The resulting treatment failure not only
Indinavir sulfate
Nelnavir mesylate
Saquinavir mesylate
Efavirenz
Lopinavir (with ritonavir)
Nevirapine
Ritonavir
Abacavir sulfate
Didanosine
Lamivudine
Stavudine
Zidovudine
Highest dose
strength (mg)
Solubility
(mg/ml)
Dose number
(Do)a
CLogP
Log P
400
250
200
200
133.3
200
100
300
200
150
40
300
1000
4.5
2.22
0.01
0.01
0.1
0.01
77
27.3
70
83
20.1
0.0016
0.22
0.36
80
53.3
8
40
0.016
0.03
0.0086
0.002
0.06
3.68
5.84
4.73
4.95
6.1
2.42
4.94
0.58
1.92
1.46
0.73
0.04
2.49
4.62
2.73
3.68
4.56
2.05
5.98
0.22
1.1
0.06
0.47
UN
67
68
P. Sharma, S. Garg / Advanced Drug Delivery Reviews xxx (2009) xxxxxx
DP
pKa
(s)
1.2
10.2
2.8
5.01
9.12
BCS class
Log P-based
CLogP-based
1
1
1
2
2
2
2
3
3
3
3
1
1
1
2
2
2
2
3
3
3
3
3
Classication criteria: dose number 1 = high solubility and N 1 = poor solubility. Estimated log P and CLogP values 1.72 and 1.35 = high permeability and b1.72 and 1.35 = low
permeability.
BCS classication: class 1 high solubility, high permeability; class 2 low solubility, high permeability; class 3 high solubility, low permeability; class 4 low solubility, low permeability.
a
Do is the ratio of drug concentration in the administered volume (250 ml) to the saturation solubility of the drug in water and calculated as Do = (Mo/Vo)/Cs where Mo is the
highest dose strength (mg), Cs is the solubility (mg/ml), and Vo = 250 ml.
Q1
ARTICLE IN PRESS
t2:26
155
156
157
158
159
160
161
162
163
164
165
166
t3:1
t3:2
t3:3
t3:4
t3:5
t3:6
t3:7
t3:8
t3:9
t3:10
t3:11
t3:12
t3:13
t3:14
t3:15
t3:16
t3:17
t3:18
t3:19
t3:20
t3:22
t3:21
t3:23
t3:24
t3:25
Capsule, liquid
Tablet, liquid
Tablet, capsule (EC), liquid
Tablet
Capsule, powder for
reconstitution
Tablet, liquid
Capsule
Tablet
60
86
3040
85
80
Nucleotide reverse
transcriptase
inhibitors (NtRTI)
Non-nucleoside
reverse transcriptase
inhibitors (NNRTI)
Protease inhibitors (PI)
Entry and fusion

inhibitors (FI)
Nevirapine
Efavirenz
Delavirdine
Etravirine
Amprenavir
Indinavir
Saquinavir
Nelnavir
Ritonavir
Atazanavir
Darunavir
Enfuvirtide
Maraviroc
Raltegravir
Integrase
inhibitors (II)
a
Abacavir
Emtricitabine
Tenofovir
Tablet, syrup
Tablet, capsule, solution
Tablet
Tablet
Capsule, solution
Capsule
Tablet, capsule
Tablet, powder
Tablet, capsule, liquid
Capsule
Tablet
Powder for subcutaneous
injection
Tablet
Tablet
83100
93
2539
N 90
4280
85
Unknown
No data
65
Erratic, 4
2080
65
No data
37
84.3
2333
No data
OF
Zidovudine
Lamivudine
Didanosine
Zalcitabine
Stavudine
RO
Nucleoside reverse
transcriptase
inhibitors (NRTI)
DP
t2:24
t2:25
F (%)a
Bioavailability.
b300 nm. In the process, the drug suspension containing macro or

micro size particles along with milling media (e.g., small spheres of
glass, zirconium oxide or polystyrene resin) is placed in an enclosed
cylindrical milling device. Milling occurs through the movement of
milling media facilitated by a rotating agitator. During the process,
mechanical energy is applied to generate sufcient strain on the solid
particles resulting in the disruption of the crystal lattice and particle
fracture. Under the stress conditions generated there is a possibility of
physical or chemical change in the crystalline solid, including
polymorphic transitions, chemical degradation, complexation or
formation of an amorphous phase and therefore, post nanosizing
samples are characterized for process induced transformation. A
TE
t2:12
t2:13
t2:14
t2:15
t2:16
t2:17
t2:18
t2:19
t2:20
t2:21
t2:22
t2:23
Dosage form
nanosized loviride formulation which was freeze-dried with sucrose

(nanopowder) did not show amorphization as indicated by powder Xray diffraction (PXRD) and Differential Scanning Calorimetry. However, there was a peak broadening in PXRD and a decrease in the Tm of
the nanosized drug, indicating the presence of nanosize crystalline
particles. Further, in a medium containing 3% sodium lauryl sulfate
(SLS) at 37 C, nanopowders showed enhanced dissolution (104%)
over (i) the freeze-dried nanosuspension without sucrose (58.1%), (ii)
the physical mixture containing sucrose (54.8%), (iii) the physical
mixture without sucrose (14.5%) and (iv) the pure loviride (64.7%)
(Fig. 1A). The increased dissolution of the nanopowder also resulted
in the enhanced transport of the drug in Caco-2 cells (Fig. 1B) as
shown by the high value of cumulative amount transported (1.59
0.02 g) compared with the physical mixture with sucrose (0.93
0.01 g) and pure loviride (0.74 0.03 g). Together, these results
indicate advantages of a nanocrystal formulation for delivery of
loviride, despite its poor aqueous solubility.
Following a similar approach, pure drug nanoparticles of a poorly
water soluble investigational anti-HIV agent (BMS-488043) were
prepared to investigate the effect on oral bioavailability in dogs [21].
BMS 488043 is a BCS class II compound with poor solubility and
optimum permeability. Moreover, the solubility is pH dependent
(0.04 mg/ml in pH 48). Nanosuspensions of this compound were
prepared using media milling. To prevent particle growth and
enhance the stability, hydroxypropylcellulose was used as the
stabilizer along with a surfactant (SLS, sodium docusate and
polyvinylpyrrolidone). The formulation was found to be stable at
4 C and room temperature for 4 weeks and the mean cumulative
particle size was 120 nm. As compared with the conventional capsule
dosage form, the nanosuspension yielded 4.7- and 4.6-fold increase in
Cmax and AUC values, respectively. This indicates that nanonization of
BMS 488043 resulted in enhanced dissolution and bioavailability.
Antiretrovirals are administered in high dose. Thus, their bioavailability is affected by the presence of food in the GI tract; food alters the
volume and composition of gastric and intestinal uids. For instance,
both lopinavir and ritonavir are water insoluble drugs and increased
uid volume and bile salt concentrations coupled with delayed gastric
emptying time lead to enhanced dissolution and bioavailability
compared to fasted state. Therefore, a lopinavir/ritonavir combination
(Kaletra capsule and oral solution) is recommended to be taken with
food. Administration of water insoluble drugs in the form of pure drug
EC
t2:9
t2:10
t2:11
Name
RR
t2:4
t2:5
t2:6
t2:7
t2:8
Class of drug
Table 3
Physicochemical properties of different protease inhibitors (PIs) (adopted from [8]).
HIV protease inhibitor
Saquinavir mesylate
Ritonavir
Indinavir sulfate
Molecular weight
767.0
CO
t2:2
t2:3
Table 2
Bioavailability of various commercially available dosage forms of antiretroviral drugs
(adapted from [63]).
Log P(o/w)a
4.1
721.0
5.2
711.9
2.9
UN
t2:1
Nelnavir mesylate
663.9
Amprenavir mesylate
601.7
4.0
(pH 7.4)
4.1
(pH 6.0)
3.3 or 4.2d
Solubility
Aq (2.22 mg/ml)
pH 7.4 (36 g/ml)
pH 6.5 (73 g/ml)
Aq (1 g/ml)
pH 7.4 (5.3 g/ml)
pH 4.0 (6.9 g/ml)
Aq (N100 mg/ml)
pH 7.4 (70 g/ml)
pH 4.8 (0.3 mg/ml)
pH 3.5 (60 mg/ml)
Aq (4.5 mg/ml)
pH 7.4 (very low)
pH 3.5 (0.5 mg/ml)
pH 2.6 (4.5 mg/ml)
Aq (0.19 mg/ml)
pH 7.4 (60 g/ml)
pH 6.8 (190 g/ml)
Papp AP to BLb (cm/s)
pKa
7.0 or 5.5
1.5 10 6
NR
3.5 10 6
pKa1 = 3.7
pKa2 = 5.9
3.0 10 6
pKa1 = 6.0c
pKa2 = 11.1c
NR
NR
NR
Aq: puried water. NR: not reported.

a
Log partition coefcient (octanol/water).
b
Apparent Caco-2 cell monolayer permeabilities in the apical to basolateral (absorptive) direction, donor concentrations range from 10 to 200 mM, the indinavir value represents
indinavir base.
c
Values reported for nelnavir base.
d
Conicting reports.
167
168
169
170
171
172
173
174
175
176
177
178
179
180
181
182
183
184
185
186
187
188
189
190
191
192
193
194
195
196
197
198
199
200
201
202
203
204
205
206
207
ARTICLE IN PRESS
4
215
216
217
218
219
220
221
222
223
224
225
226
227
228
229
230
231
232
233
234
235
236
237
238
239
240
241
242
243
244
245
246
247
248
249
250
251
252
253
OF
3. Polymer based nanotechnologies
285
3.1. Polymeric micelles
286
Polymeric micelles are nanostructures of b100 nm diameter that

have been utilized for improving aqueous solubility, intestinal
permeability and disease site targeting of several drug molecules.
Compared to the conventional surfactant based micelles, polymeric
micelles are composed of block copolymers. Although the structural
coreshell arrangement is similar to surfactant based micelles,
polymer micelles self associate at much lower concentration (low
critical micelle concentration, CMC), typically 10 610 7 M compared with 10 310 4 M for surfactant based micelles [30,31].
Consequently the thermodynamic and in vivo stability of polymeric
287
RO
254
DP
213
214
rst-pass metabolism and results in dose reduction which is highly

signicant considering the adverse effects associated with the anti-HIV
drugs.
It is also worth mentioning that media milling, developed by
Liversidge et al., is a proprietary wet milling technology (Nanocrystal)
of the Elan Corporation [12,27] and there are at least four oral dosage
forms in the market containing pure drug nanoparticles; i.e., Rapamune (Wyeth), Emend (Merck), Tricor (Abbott Laboratories) and
Megace ES (Bristol-Myers Squibb) [28]. Thus, Nanocrystal is a
validated technology with well established process controls and scale
up feasibility. The choice of stabilizer during media milling signicantly
affects the nal size of the particles. The process of milling generates
numerous nanosize particles with a very high surface area. An increase
in the surface area (A) is associated with an increase in the surface
free energy (G) by the equation G = A s/l where, s/l is the
interfacial tension between the surface of solid and the surrounding
liquid phase. In order to reduce the free energy, particles tend to
agglomerate resulting in an increased particle size and reduced
surface area. Under these circumstances, inclusion of a surface
stabilizer leads to a decrease in the G by decreasing the interfacial
tension s/l. The overall effect of the addition of a stabilizer is
reduction in the particle growth, thereby conferring physical
stabilization to the nanosuspension [29]. Further, because the
benets of nanosized particles (such as high surface area) can be
exploited only by keeping particles dispersed, it is possible to convert
nanosuspension obtained after media milling to solid products for
stability enhancement and protection against aggregation using
processes such as spray drying, lyophillization and wet granulation
following media milling. Such solid products can be powder for
reconstitution, pellets or powders for capsule lling or tablet
compression which ultimately improves patient compliance.
TE
211
212
nanoparticles reduces the bioavailability variability brought by the

presence or absence of food [22,23]. However, to elicit this effect,
nanoparticles should re-disperse homogeneously in the GI uids after
administration. Further, as expected, this is a dose dependent effect
and at higher doses of poorly water soluble drugs some variation may
be observed in fed and fasted state bioavailability [22].
As an i.v. formulation, the major advantage of nanosuspension is
the avoidance of precipitation following administration. Since the
formulation contains a poorly water soluble drug as nanosized
particles, their biodistribution depends upon the solubility of the
drug in the blood [24]. After administration, if the particles dissolve
rapidly due to dilution in the blood circulation, the biodistribution will
be similar to that of the solution formulation. However, in case of slow
dissolving particles, macrophage uptake occurs. Other advantages of
nanosuspension as i.v. formulation include (i) high dose per volume
(high drug loading), thus, requiring low volumes for injection and
(ii) reduced toxicity due to elimination of co-solvents and solubilising
agents (such as cremophor) along with scale up feasibility.
In a proof of concept study, Dou et al. demonstrated the
concentration-dependent in vitro uptake of indinavir nanocrystals
(mean particle size 1.6 m, stabilized with phospholipid stabilizer) in
monocyte-derived macrophages [25]. Following incubation, 50%
macrophages contained nanocrystals after 2 h, which increased to
95% after 8 h. The authors reported that after cellular uptake, indinavir
nanocrystals were exposed to the low pH of phagolysosomes,
resulting in dissolution and release of indinavir in the extracellular
matrix. Furthermore, incubation of HIV infected macrophages with
indinavir nanocrystals showed signicantly reduced reverse transcriptase activity and expression of HIV-1p24 antigens compared with
soluble drug in culture uids. Considering that monocyte-derived
macrophages are HIV reservoirs, results of this study indicated the
viability of transporting nanocrystals engulfed by the macrophages
across the blood brain barrier.
Nanosuspensions can also be used as a long acting parenteral
formulation, as reported recently for rilpivirine (TMC278) [26]. A long
acting formulation has a major advantage of minimizing the dosing
frequency, thereby, increasing compliance to the HIV therapy. Rilpivirine is a potent NNRTI and is poorly soluble in both water and oil. After
intramuscular (i.m.) and subcutaneous (s.c.) administration of a single
dose of 5 mg/kg of 200 nm size particles in dogs, a sustained plasma
concentration was achieved up to 3 months. Compared with i.m., s.c.
administration resulted in sustained release of rilpivirine (Cmax, IM:
173 ng/ml and Tmax, IM: 24 h versus Cmax,SC: 38 ng/ml and Tmax, SC:
144 h). This recent study is the rst to provide a proof of concept for a
nanosuspension as a long acting formulation. In addition to minimizing
the dose frequency, the parenteral route of administration also avoids
RR
EC
209
210
CO
208
Fig. 1. (A) Dissolution proles: freeze-dried nanosuspension without sucrose (), physical mixture without sucrose (), nanopowder (), physical mixture with sucrose (),
untreated loviride (). Dissolution of the nanopowders is complete within minutes. (B) Cumulative transported amount of loviride as a function of time in Caco-2 experiments:
nanopowder (), physical mixture (), untreated loviride () (adopted from [20]).
UN
Q13
255
256
257
258
259
260
261
262
263
264
265
266
267
268
269
270
271
272
273
274
275
276
277
278
279
280
281
282
283
284
288
289
290
291
292
293
294
295
296
ARTICLE IN PRESS
312
313
314
315
316
317
318
319
320
321
322
323
324
325
326
327
328
329
330
331
332
333
334
335
336
337
338
339
340
341
342
343
344
345
346
347
348
349
350
351
352
353
354
355
356
357
358
359
360
361
362
OF
310
311
RO
308
309
DP
306
307
TE
304
305
363
364
365
366
367
368
369
370
371
372
373
374
375
376
377
378
379
380
381
382
383
384
385
386
387
388
389
390
391
392
393
394
3.2. Polymeric nanoparticles
395
Polymeric nanoparticles (NPs) are b1000 nm in size and are

composed of biodegradable or biostable polymers and copolymers.
The active agent can be (i) entrapped or encapsulated within the
particle, (ii) physically adsorbed on the surface, or (iii) chemically
linked to the surface of the particle. There are many studies where NPs
have been explored for the delivery of anti-HIV molecules [5761].
Recently Govender et al. and Ojewole et al. have reviewed this subject
396
397
EC
303
RR
301
302
0.001% and 0.01% showed signicant reduction in HIV replication.

When compared with control (no treatment), HIV replication was
reduced to 55% and 44% at 0.001 and 0.01% concentrations. The
activity was further enhanced in the presence of nelnavir compared
to either P85 or nelnavir alone (Fig. 2). In vivo, the antiretroviral
combination without P85 (0.2%) reduced the HIV-1 p24 positive cells
to 35%. P85 alone and in combination with the antiretroviral showed
greater effect, 13.4 and 7.0% respectively, compared to the control
(68.5%) (Fig. 3). This inhibition of the HIV replication by P85 alone
could be due to its direct interaction with P-gp or glycolipid
membrane of HIV [55] which results in membrane disruption. In a
recent study, inuence of P85 on the interaction of nelnavir and
saquinavir with P-gp expressed on MDR1-transfected epithelial
MadinDarby canine kidney (MDCKII) and MDR1-transfected porcine
kidney epithelial cells (LLC-PK1) cells was investigated [36]. P-gp
utilizes the energy released from the hydrolysis of ATP. This action is
mediated by ATPase enzyme [56]. PIs have been shown to stimulate
the ATPase activity leading to their efux. In the study, P85 was found
to inhibit the basal activity of ATPase, thereby, inhibiting the efux
action of P-gp. Moroever, P85 (0.01%) was also found to inhibit MRP1
and MRP2, in addition to P-gp, leading to enhanced accumulation of
saquinavir in both MDCKII and LLC-PK1 wild type and MDR1 cells
[36].
Comparative studies of P85 with other Pluronic (F 127, F 68) have
reected the importance of structural requirements of PEOPPOPEO
units for the biological effects; molecules having intermediate length
of PPO and hydrophobicity, such as P85, being more effective [36,56].
For oral delivery of antiretroviral molecules, for example saquinavir,
whose absorption is limited by the efux action of multidrug
transporters, inclusion of P85 in the formulation, at a concentration
less than the expected CMC after dilution in GI uid, may lead to
enhanced bioavailability.
CO
299
300
micelles is relatively high [16,32]. Polymeric micelles consist of a

hydrophobic block which constitutes the core and a hydrophilic block
constituting the shell of the micelle. This coreshell arrangement
facilitates their utilization as a drug delivery nanocarrier where
depending upon the polarity the drug molecule can be entrapped in
the (i) core (non polar molecule), (ii) shell (polar molecule) and
(iii) in-between core and shell (intermediate polarity). As a result,
micelles are suitable for (i) solubilization of poorly water soluble
drugs, (ii) protection against chemical degradation and metabolism
and (iii) controlled release. Additionally, surface properties of micelles
can be tailored by attaching hydrophilic blocks to antibodies or other
ligands specic for the type of receptors present on the disease site.
For example, micelles of polyethylene (PEG)polylactide copolymer
with a surface modied with galactose and lactose units specically
interact with lectins [33]. Lectin receptors are present on HIV viral
reservoirs such as T lymphocytes, dendritic cells and macrophages,
and therefore, this can be a promising approach for viral reservoir
targeting.
The use of polymeric micelles for oral delivery has been recently
discussed by Bromberg [32]. Various polymers have been used as
hydrophilic shell forming blocks in polymeric micelles such as PEG
(molecular weight 1 to 15 kDa), poly(N-vinyl-2-pyrrolidone) (PVP),
poly(vinyl alcohol) (PVA), polyethyleneimine and poly(ethylene
oxide) (PEO). Examples of hydrophobic core forming blocks include
L-lysine, aspartic acid, caprolactone, D,L-Lactic acid, propylene oxide
and others [16,34]. Studies have been reported where polymeric
micelles composed of different block copolymers were used for oral
delivery [35]. Among these, block copolymers of poly(ethylene
oxide)poly(propylene oxide) (PEOPPOPEO), also known as Pluronics, have been used to investigate their potential as intestinal
permeability enhancers of antiretroviral drugs [36]. Pluronic, P85
modulates the activity of ATP binding cassette transporters [3639]
that have been shown to reduce the oral bioavailability [4045] and
CNS permeability [10,4547] of many PIs and NRTIs. For example, the
P-gp mediated efux transport of the PI saquinavir has been reported
to be a key factor for its exceptionally low bioavailability [44].
Similarly, among other reasons such as poor water solubility, CYP3A4
metabolism and systemic clearance, P-gp mediated efux has also
been reported to play a role in the low bioavailability of indinavir,
atazanavir and lopinavir; concomitant administration of a low dose Pgp and CYP 3A4 inhibitor, ritonavir (also a PI), results in their
enhanced bioavailability [4852]. Inhibition of P-gp by P85 was rst
observed in SKVLB multidrug resistant (MDR) cancer cells where 0.01
to 1% P85 concentration resulted in 700 times more cytotoxicity (or
MDR reversal) of otherwise MDR drug daunorubicin [53]. In addition,
since the effect was also observed at the 0.01% concentration at which
P85 was present as unimers, it was proposed that the observed effect
on P-gp was not due to micelles. Batrakova et al. showed a similar
effect of P85 on Caco-2 monolayers (in vitro model for intestinal
epithelium) over-expressing P-gp [54]. Below CMC, presence of P85
unimers caused a signicant uptake and efux inhibition of P-gp
dependent probe rhodamine 123 in Caco-2 cells, whereas, the effect
observed with other nonionic detergents, Cremophor EL and Tween
60 was signicantly lower.
The effect of different P85 concentrations (0.01 to 1%) on the
permeability of the NRTI AZT in bovine brain microvessel endothelial
cells (BBMEC, in vitro model for blood brain barrier) and Caco-2 cell
was studied among other drugs. At all P85 concentrations, AZT
permeability enhancement was signicant due to P-gp and MRP
mediated efux inhibition [37]. Further, the effect of P85 mediated
inhibition of P-gp efux on the efcacy of antiretrovirals was assessed
[55]. P-gp inhibition by P85 could enhance the in vitro efcacy of
nelnavir in HIV-1 infected monocyte-derived macrophages and the
in vivo efcacy of AZT, lamivudine (3TC), and nelnavir combination
therapy in HIV 1 encephalitis (HIVE) severely combined immunodeciency (SCID) mice. Interestingly, in the in vitro studies P85 alone at
UN
297
298
Fig. 2. Pluronic block copolymer P85 inhibits HIV-1 replication in monocyte-derived

macrophages (MDM) (adopted from [55]). Effect of combined treatment with
(nelnavir) NEL (0.01 and 0.001 mol/L) and P85 (0.01% and 0.001%) on HIV-1
replication in MDM as detected by reverse transcriptase (RT) activity at day 12,
normalized to control cells. P b 0.01, # P b 0.05 NEL + P85 as compared with
NEL (0.01 mol/L) or P85 (0.01%) alone. Results are presented as mean standard
error (SE).
398
399
400
401
402
ARTICLE IN PRESS
411
412
413
414
415
416
417
418
419
420
421
422
423
424
425
426
427
428
429
430
431
432
433
434
435
436
437
438
439
440
441
OF
TE
409
410
RR
EC
407
408
CO
405
406
[62,63]. In most cases, polymeric nanoparticles have been used to

target antiretrovirals to (i) macrophages/monocytes [59,60,64,65]
and (ii) CNS [57,58,61] which act as viral reservoir sites during HIV
infection [6669]. Macrophages have been reported to be a major
cause of dissemination of the infection in the body [69] in the later
stages of the disease during which there is a continuous depletion of
CD4+ T lymphocytes. During this period, virus production from these
mature non-proliferating macrophages/ monocytes is dramatically
enhanced without being affected by the lethal effect of the replicating
virus.
Nanoparticulate mediated targeting of macrophages is well known
and has been reported by several authors [64,7072]. Following i.v.
administration, nanoparticles are removed from the blood circulation
by macrophages [73,74]. The recognition of particles by macrophages
is mediated by a process called opsonization [75,76]. When the
distance between the particles and the opsonins is sufciently small,
they can bind to the surface of particles by any of the interactions such
as van der Waals, electrostatic, ionic etc. After binding to the surface,
particles become recognizable by macrophages and phagocytosis
takes place [75]. Size and surface characteristics of nanoparticles are
the determinants of their clearance kinetics and tissue distribution
[73,74,76]. These properties interfere with the opsonization process
and affect the macrophage uptake.
In 1992, Schaffer et al. showed the monocyte/macrophage
targeting of nanoparticles using macrophages separated from HIV
infected individuals [64]. In addition, these authors also showed the
effect of particle size, concentration and surface characteristics on the
macrophage uptake. Subsequently, other studies also reported the
macrophage targeting of nanoparticles [65,77].
HIV infection of brain macrophages, i.e. perivascular macrophages,
meningeal macrophages, the macrophages of choroid-plexus and
microglial cells, is directly linked to HIV associated dementia complex
(HAD) [9]. Several works reported on the targeting of entrapped or
conjugated drugs to the CNS. For example, Zensi et al. recently studied
the brain uptake of human serum albumin nanoparticles coated with
covalently bound apolipoprotein E after i.v. administration in mice
[61]. The presence of apolipoprotein bound nanoparticles in the
endothelial cells of the brain capillaries and neurons demonstrated
their preferential brain uptake over non-apolipoprotein bound
UN
403
404
RO
Fig. 3. Effect of P85 (0.2%) and triple drug combination (antiretroviral-ART) on viral
replication in HIVE SCID mice (adopted from [55]). At day 7, the combined ART-P85 and
ART alone groups showed a signicant decrease of HIV-1 p24 expressing MDM (9% and
13%, respectively) while the P85 alone was not different from control. At day 14, most
notable effects were seen in the P85 alone and ART-P85 groups (6% to 15% of HIV-1 p24positive MDM), which were superior to the ART group (35% p24-positive cells). Bar
values represent mean SE.
nanoparticles. In addition, transportation of apolipoprotein bound

nanoparticles through tight junctions of the endothelial cells and the
luminal membrane was not observed, and therefore, a transcytosis
mechanism of transport was suggested for the internalization. An
extensive discussion of this application is out of the scope of the
review and it is covered by another review of this Theme Issue.
Biodegradable polymeric nanoparticles have been used for the
vaginal delivery of unconventional anti-HIV agents such as chemokines [77] and siRNA (small interfering ribose nucleic acid) [78]. The
aim behind the development of a vaginal formulation (known as
microbicide) is to prevent transmission of HIV during sexual
intercourse. A microbicide formulation is intended for intravaginal
application before sexual exposure. The effort needed in this eld of
research is also supported by the fact that nearly half of the HIV
infected population is female, and majority of them acquire the
infection during intercourse. The major advantage of using nanoparticles includes protection of the actives against degradation in the
vaginal environment. Also, nanoparticulate delivery may assist by
direct action on one or more of the target sites such as submucosal
epithelium (Fig. 4). Nanoparticles have been reported to permeate
through the interstitial spaces in tissue due to their nanosize which
may result in enhanced cellular uptake [79]. Recently, Lai et al. have
shown that physicochemical properties of polyethyleneglycol (PEG)conjugated nanoparticles (carboxyl-modied polystyrene) such as
size, surface charge and particle concentration signicantly affect
their diffusion through cervico-vaginal mucus [80]. Specically, it was
reported that 200500 nm size particles (b0.01% by weight) linked
with 2 kDa PEG diffuse through the mucus faster than 100 nm size
particles. This could be considered as signicant because an effective
microbicide should act on the luminal as well as basal side of the
vaginal epithelium. In order to reach the basal side, traversing through
mucus is a prerequisite for nanoparticles. Further, Cu et al. have
reported similar results of higher diffusion rate of PEG modied
(copolymer polylactic acid-co-glycolic acid) nanoparticles [81]. The
diffusion rate was dependent upon PEG type and concentration with
rates 10 times higher compared with non-PEG modied nanoparticles. In addition to the prophylactic application, these ndings also
widen the nanoparticle horizons towards diagnostic applications in
HIV.
Compliance of some protease inhibitors (ritonavir, indinavir) in
pediatric population is low due to the poor palatability (extreme
bitterness) of the liquid formulation [52]. In order to mask the
bitterness of indinavir sulfate, Chiappetta et al. [82] prepared pH
sensitive microparticles of Eudragit E 100 which is a cationic
copolymer consisting of dimethylaminoethyl methacrylate and
neutral methacrylic esters. Three different microparticle formulationsMP20, MP40 and MP60containing three different theoretical
indinavir loadings, 20%, 40% and 60%, respectively, were prepared. For
MP20 and MP40, 80% (by weight) of the particles were between 105
and 210 m, whereas, for MP60 the mean particle size was higher
(210420 m) with broad distribution. The in vitro release studies of
all the formulations showed 100% drug release in 5 min at acidic pH
1.5; in contrast, at pH 6.8 slow release of indinavir from the
microparticles was observed (Fig. 5). This shows that in saliva like
pH conditions (pH 6.8), the polymer remained unprotonated resulting
in poor solubility, and therefore, most of the bitter drug remained
inside the microparticles. However, following exposure to the acidic
pH (gastric pH), indinavir was immediately released due to the
solubilization of polymer matrix mediated through protonation of the
amine side chains of the polymer. When the MP20 and MP40
microparticles were dispersed in tap water (simulation of extemporaneous reconstitution), indinavir concentrations after 1 min were
within the threshold limits (2045 g/ml) that are considered
palatable. However, MP60 produced higher indinavir concentrations
(1215 g/ml) which were unpalatable. Moreover, after 5 min compared with MP40 and MP60, only MP20 produced palatable indinavir
DP
442
443
444
445
446
447
448
449
450
451
452
453
454
455
456Q3
457
458
459
460
461
462
463
464
465
466
467
468
469Q4
470
471
472
473
474
475
476
477
478
479
480
481
482
483
484
485
486
487
488
489Q5
490
491
492
493
494
495
496
497
498
499
500
501
502
503
504
505
506
507
ARTICLE IN PRESS
7
RO
OF
515
516
517
518
3.3. Dendrimers
519
A dendrimer is a polymeric nanostructure (b100 nm) built around

a core unit. There are several branching units around the core units in
a layer by layer fashion which denes the growth, size and the
microenvironment within the dendrimer [83]. Dendrons are formed
after removal of core units and can be divided into (empty) core, the
interior (branching units) and the periphery (end groups). The empty
core of dendrons can be utilized for the entrapment of drug molecules
for solubilization, controlled release, targeting or protection from
surrounding degrading environment. Dendrimers offer unique properties such as uniform particle size (especially for lower generation i.e.
up to G-3), poly-valency of the end groups which helps in binding to
diverse receptors and an ability to bind a variety of targeting agents to
their high density peripheral functional groups.
The potential of the oral delivery of dendrimer has been explored
with cationic and anionic PAMAM dendrimers [8490]. Various
reports have indicated that the permeability of PAMAM dendrimers
across Caco-2 monolayers and everted rat intestinal sac is dependent
on their size, surface charge, charge concentration and surface
modication [91]. Additionally, conjugation of propranolol (a P-gp
susbtrate) to G-3 PAMAM dendrimer was shown to bypass the efux
action of P-gp [92]. The proposed mechanisms for the dendrimer
passage across the GI epithelium are energy-dependent adsorptive
mediated endocytosis and paracellular pathway [91]. Wiwattanapatapee et al. studied the tissue uptake and serosal transport of 125I
labelled rst and half generation PAMAM dendrimers across the
everted rat intestinal sac [93]. Rate of tissue uptake for G-5.5 was
signicantly higher than lower generations G-2.5 and 3.5 whereas the
rate of serosal transfer was relatively similar between the three
generations. Moreover, contrary to anionic dendrimers, the rate of
tissue uptake of G-3 and G-4 cationic dendrimers was higher than
serosal transfer and both showed similar rates for tissue and serosal
520
521
EC
514
RR
512
513
CO
510
511
concentrations. Taste masking studies were conducted on healthy

human volunteers using indinavir solutions having concentrations
equivalent to that produced after keeping the formulations in tap
water for 1 and 5 min. MP20 scores were within the limits (b2) set for
palatability. However, MP40 and MP60 formulations scored more
than 2 and, therefore, were unpalatable. These results indicated that
microparticles of Eudragit E100 polymer could efciently entrap 20%
indinavir (theoretical loading) with acceptable palatability. Even
though this work focused on microparticles, the results of this study
showed the utility of a polymer based technology in improving the
palatability of indinavir formulations.
UN
508
509
TE
DP
Fig. 4. Mode of transmission of human immunodeciency virus (HIV) through female genital mucus and target sites for microbicides (adapted from [109]). HIV can enter genital
tract through submucosal tissue (consisting of squamous stratied epithelium (left) and cervical columnar epithelium (right)). Transmission of HIV across the submucosal tissue can
occur through various pathways [110]: following entrapment in mucus (A) and diffusion towards single layer of columnar cells through which endocytosis (B) can occur. Langerhans
cells present in stratied epithelium may also get infected with virus (C). Virus can permeate through ulceration or lesions in the stratied epithelium (D). Following entry through
these pathways, HIV can infect T cells (E), macrophages (F) and dendritic cells (G). Through these cells, HIV can travel via lymphatics to regional lymph nodes leading to further viral
dissemination in the body. To effectively counter HIV infection, microbicides must be effective against these multiple sites shown above.
Fig. 5. Released indinavir (expressed as %) versus time for MP20 and MP40 systems
(adopted from [104]). Microparticles (100 mg) were dispersed in the test medium
(500 mL, pH values 1.5 and 6.8) and assayed in a dissolution test using the USP
dissolution apparatus 2, at 37 C.
522
523
524
525
526
527
528
529
530
531
532
533
534
535
536
537
538
539
540
541
542
543
544
545
546
547
548
549
550
ARTICLE IN PRESS
562
563
564
565
566
567
568
569
570
571
572
573
574
575
576
577
578
579
580
581
582
583
584
585
586
587
588
589
590
591
592
593
594
595
596
597
598
599
600
601
602
603
604
605
606
607
608
609
610
611
612
613
614
615
616
OF
560
561
RO
558
559
TE
557
results were corroborated with confocal microscopy, where uorochrome-labeled siRNA either naked or complexed with dendrimer
were observed in the interior of the cells of lymphocytic cell line
SupT1 (Fig. 7). Contrary to SupT1 cells, the transfection efciency of
siRNA alone was poor ( 1%) in HIV infected peripheral blood
mononuclear cells (PBMC) after 3 h; however, following complexation with 2G-NN16 the dendriplex showed signicantly higher
(40%) transfection efciency. After 24 h, the transfection efciencies
of siRNA and dendriplexes were more or less similar with minimal
cytotoxicity (80% cell viability) (Fig. 8). In addition, dendriplexes
were efcient in reducing HIV replication in SupT1 and PBMC.
Compared with siNEF and siCocktail (containing siP24, siNEF and
siGAG1) alone, treatment with 2G-NN16 dendriplexes showed 40%
HIV inhibition for +/ charge ratio of 2 (Fig. 9). The low cytotoxicity
coupled with high transfection efciency and stability of siRNA
provided by carbosilane dendrimer demonstrates the importance of
their development as a delivery system for siRNA.
Dutta et al. prepared G-5 PPI dendrimers for targeting of efavirenz
(EFV) [106] and lamivudine (3TC) [107] to human monocytes/
macrophages and MT-2 cells, respectively. For EFV targeting, the C
terminus of a tetrapeptide (ThrLysProArg)tuftsin (tu)was
conjugated to the amino end group of the PPI. Tetrapeptide tu binds
specically to the macrophages/monocytes and polymorphonculear
nucleocytes and also activates the immune system. EFV loading was
found to be 37.4% in PPI and 49.3% in tuPPI dendrimers. The higher
drug loading efciency for tuPPI was attributed to the availability of
more functional groups for complexation with EFV. The presence of tu
also signicantly delayed the release of EFV from the dendrimer
(144 h for tuPPI as compared with 24 h for PPI) due to the presence
of dense steric groups of tu at the surface of dendrimer. Presence of
bulky tu also shielded the positive charges of the PPI dendrimer and
led to a signicantly decreased cytotoxicty in macrophage/monocyte
cells compared with the unmodied PPI dendrimer. The cellular
uptake of EFV was signicantly (34.5 times) higher when present in
tuPPI dendrimer compared with free drug which was attributed to
the targeting effect of tuftsin present on the periphery of PPI
dendrimer. Moreover, HIV infected macrophages/monocytes showed
higher uptake of tuPPI as compared to uninfected cells possibly due
to their activated state. tuPPI loaded with or without EFV also led to
inhibition of HIV growth at low concentrations as determined by p24
antigen assay.
For the targeting of 3TC, Dutta et al. prepared G-5 mannosylated
PPI dendrimers (MPPI) by divergent synthesis [107]. The aim was to
target lectin receptors (molecular target for sugar molecules such as
mannose). Mannose receptor is a transmembrane glycoprotein
expressed by macrophages and has the ability to phagocytose
nanoparticles coated with saccharides [70]. This has been reported
for mannan-coated nanoparticles which show enhanced uptake in
mannose receptor-positive mouse macrophage cell line (J-774E)
[108]. Puried mannose receptor shows specic interaction with
monosaccharide ligands such as mannose and fucose [70]. Compared
with PPI dendrimer, there was an increase in the drug entrapment
efciency from 35.7 to 43.2% for MPPI due to the steric hindrance
provided by the presence of mannose on the surface of PPI end groups.
Cytotoxicity studies of blank dendrimer in MT2 cell line showed
decreased toxicity for MPPI (0.156 nM/ml) compared with PPI
(0.039 nM/ml) due to shielding of the positive charges of PPI by
mannose. Cellular uptake studies in MT2 cells showed 21 and 8.3
times higher uptake for MPPI compared to free 3TC and PPI,
respectively, indicating ligand specic internalization of MPPI dendrimers. Furthermore, MPPI dendrimers containing 3TC also showed
superior anti-HIV activity (determined by p24 antigen assay)
compared with free 3TC and PPI containing 3TC. The anti-HIV activity
of MPPI dendrimers at 0.156 ng/ml was 1.5 fold higher than free 3TC
at 0.625 ng/ml suggesting superior targeting ability of MPPI
dendrimers.
RR
EC
555
556
CO
553
554
transfer for the rst 60 min [93]. Also, anionic PAMAM dendrimers
have shown to be more permeable in Caco-2 monolayers compared to
cationic PAMAM dendrimers [84,89]. The success of dendrimers as
delivery systems lies in their biocompatibility and acceptability.
PAMAM and PPI dendrimers are non-biodegradable which after
chronic administration can cause unpredictable toxicity. Therefore,
the size of the dendrimers must be controlled to facilitate renal or
hepatic clearance [94]. Dendrimers with cationic (amino) end groups
such as PAMAM and PPI have been shown to cause concentrationdependent toxicity and hemolysis, whereas, neutral or anionic
dendrimers are less toxic and less hemolytic [82,94]. In addition,
modication of the terminal amino surface groups with anionic or
neutral groups (for example PEG 2000) reduces the toxicity
[84,95,96]. Results of in vitro studies correlate well with in vivo
studies and intraperitoneal administration of doses above 10 mg/kg of
cationic melamine dendrimers caused liver toxicity [97]. Administration of a 160 mg/kg dose caused 100% mortality within 12 h. However,
for a structurally similar dendrimer, modication of the terminal
cationic groups with neutral polyethylene oxide reduces the toxicity
and i.v. or intraperitoneal administration of doses more than 1 g/kg
produced no toxic effects [98].
Interfering ribonucleic acid (siRNA) are small pieces of double
stranded RNA used to halt gene expression by participating in RNA
interference pathway. Such gene silencing activity is sequence specic
and results in degradation of messenger RNA complementary to
siRNA. Noticeably, many siRNA based therapeutics have recently
entered clinical trials [99,100] and are promising approaches for
silencing the genes involved in pathological conditions. Studies have
shown these approaches to be effective against HIV [101,102].
However, the in vivo stability of siRNA is poor and it undergoes
degradation under physiological conditions. Moreover, poor cellular
uptake and transfection efciency coupled with endosomal degradation are the major drawbacks associated with siRNA delivery. Being
polyanionic, complexation of siRNA to polycationic biodegradable
polymers is a promising approach for efcient delivery [104]. Weber
et al. developed dendriplexes of positively charged carbosilane
dendrimer and short negatively-charged siRNA [105]. The G-2
ammonium-terminated carbosilane dendrimers were 2G-NN8 (2G
CBS-(OCH2CH2NMeCH2CH2N+Me+
3 I )8) and 2G-NN16 (2G-CBS(OCH2CH2N+Me2CH2CH2N+Me3I)8) carrying 8 and 16 positive
charges, respectively. These dendrimers were water soluble, efcient
in binding siRNA and able to release their load in a time dependent
manner as a result of carbon-silicon bond hydrolysis [105]. Specic
polyanionic siRNA sense and antisense sequences coding for P24,
GAG1 and NEF HIV proteins were used for binding to dendrimers.
Heparin competition assay showed that compared to 2G-NN16, siRNA
were strongly bound to 2G-NN8. Moreover, after addition of 2G-NN16
to 500 nM siGAG1 zeta potential increased from 20 to + 5 for +/
charge ratio 1 up to 3, with a further increase in +/ charge ratio
resulting in a gradual increase in zeta potential as well as size (Fig. 6).
For charge ratios up to 2, the measurements showed a negative zeta
potential and size b300 nm. Results of agarose gel electrophoresis
showed that attachment of siRNA to the quaternary ammonium
groups of carobosilane dendrimer increased the stability of siRNA
against RNase-mediated degradation. Further, tests performed to
measure the membrane rupture, cell viability, metabolic activity and
cell proliferation showed decreased cytotoxicity of the dendriplex in
lymphocytic cell line SupT1 compared with dendrimer alone because
of the shielding of the positive charges of the dendriplex which were
otherwise exposed in the absence of siRNA in the dendrimers. The
cytotoxicity was dependent on +/ charge ratios of dendriplexes,
with charge ratios b8 being non-toxic. Similarly, results of ow
cytometry performed with a uorochrome-labeled siRNA either alone
or in dendriplexes showed that the transfection efciency of the
dendriplexes was charge-dependent and highest for either siRNA
alone or with dendriplexes with +/ charge ratio of 1 and 2. These
UN
551
552
DP
617
618
619
620
621
622
623
624
625
626
627
628
629
630
631
632
633
634
635
636
637
638
639
640
641
642
643
644
645
646
647
648
649
650
651
652
653
654
655
656
657
658
659
660
661
662
663
664
665
666
667
668
669
670
671
672
673
674
675
676
677
678
679
680
681
682
ARTICLE IN PRESS
9
OF
4. Conclusion and future directions
684
The growing number of HIV infected individuals and related

deaths reects upon the efforts required to tackle this pandemic. To
date there are over twenty ve drugs approved for the treatment. The
effect of antiretroviral therapy on the quality of life of HIV patients has
been enormously positive; however, toxicity, adverse drug reactions,
suboptimal bioavailability due to poor physicochemical properties,
TE
EC
RR
689
CO
687
688
UN
685
686
impaired biodistribution in HIV reservoirs, emergence of drug

resistance, requirement of drug monitoring and lifelong adherence
are the problems associated with antiretroviral treatment.
The studies discussed in this review provide the proof of concept
that rational use of nanoscale drug delivery systems has the potential
to tackle the water solubility, stability, bioavailability and viral
targeting issues associated with antiretroviral therapy. Most of these
studies have utilized cell cultures for in vitro and small animals for in
DP
683
RO
Fig. 6. Zeta potential and particle size of dendriplex. (A) Zeta potential of 500 nM siGAG1 upon addition of 2G-NN16 at varying +/ charge ratios. (B) Particle size (estimated by
intensity) of the same dendriplex formulations. Dendriplex formed in 0.15 mM HEPES buffer, pH 7.4, 25 C (adopted from [105]).
Fig. 7. Complete internalization of siRNA into T cells (adopted from [105]). Confocal microscopy images of SupT1 cells after 20 h incubation with mock treatment (a) or with Cy3labeled siRNA (red) alone (b) or complexed with 2G-NN16 at a +/ charge ratio of 1 (c) or 2 (d). Cell membranes are labeled with CD45-FITC antibodies (green). Percentages of
positive cells for siRNA uptake are indicated. (For interpretation of the references to colour in this gure legend, the reader is referred to the web version of this article.)
690
691
692
693
694
695
696
697
ARTICLE IN PRESS
vivo studies. With the establishment of the proof of concept, there is a

need to test efcacy of these delivery systems in higher animal models
such as rhesus macaque monkeys, which are routinely used in the HIV
drug development. The unavailability of references on the efcacy of
nanoscale delivery system in higher animal models and humans
indicates that the research in this eld is still in the early stages.
A major factor that would govern the applicability of the nanoscale
drug delivery systems is the cost of the treatment compared to the
standard antiretroviral therapy. Development of nanoscale delivery
systems frequently requires specialty chemicals and unique manufacturing procedures which signicantly add to the cost of the nal
product. In addition, complexities involved in scale up and quality
control of the nal formulation need to be considered in advance.
Majority of the studies published in the literature have focused on
developing a drug delivery system suitable for a single drug only.
Although, most of the antiretroviral drugs can reduce the viral load
and increase CD4 count individually, to sustain the levels and avoid
drug resistance, a combination of three or more drugs is required.
Therefore, further studies on nanoscale delivery systems should focus
on combinational therapy. The loading efciency of a nanoparticulate
system would dictate the size and route of administration of the
dosage form. For example, for a 500 mg dose, 15% loading efciency
would require 3.3 g of the nanoparticulate system to be administered.
Such an amount can only be administered through oral route. In the
absence of HIV eradication approach, the major challenge associated
with current treatment is poor patient compliance. In this regard,
development of a cost effective dosage form which can be selfadministered would be preferred.
The advantages associated with nanoscale drug delivery systems
are enormous. At the same time, one has to judiciously consider the
practicality of the approach within clinical settings. In this context,
insights gained from the application of nanoscale drug delivery
systems in other diseases, such as cancer, will be useful in assisting the
rational development of delivery systems for anti-HIV drugs. With a
number of challenges associated with HIV prophylaxis and treatment,
there is an ever-growing need for the development of new drug
delivery systems. Nanotechnology based drug delivery systems hold
promise to alleviate the problems associated with anti-HIV drugs and
improve the quality of life of the HIV infected patients.
698
699
5. Uncited reference
737Q7
TE
DP
RO
OF
10
UN
CO
RR
EC
Fig. 8. Transfection and toxicity proles of dendriplexes on HIV infected T cells

(adopted from [105]). Dendriplex at varying +/ charge ratios after 3 h incubation (A)
and 24 h incubation (B). Toxicity was measured by LDH enzyme release, while
transfection and cell viability were measured by ow cytometry. Lipofectin was used
as a comparative control for both toxicity and transfection.
Fig. 9. Sequence specic inhibition of HIV in PBMC after 24 h incubation with

dendriplexes at varying +/ charge ratios with siNEF (black bars) and siCOCKTAIL
(grey bars). Toxicity values as measured by LDH enzyme release after 3 h incubation are
shown (white bars). Data are shown as mean SEM; n = 3 (adopted from [105]).
700
701
702
703
704
705
706
707
708
709
710
711
712
713
714
715Q6
716
717
718
719
720
721
722
723
724
725
726
727
728
729
730
731
732
733
734
735
736
[103]
738
References
739
[1] E. De Clercq, Anti-HIV drugs: 25 compounds approved within 25 years after the
discovery of HIV, Int. J. Antimicrob. Agents 33 (2009) 307320.
[2] D.D. Richman, D.M. Margolis, M. Delaney, W.C. Greene, D. Hazuda, R.J. Pomerantz,
The challenge of nding a cure for HIV infection, Science 323 (2009) 13041307.
[3] C. Flexner, HIV drug development: the next 25 years, Nat. Rev. Drug Discov. 6
(2007) 959966.
[4] G.L. Amidon, H. Lennernas, V.P. Shah, J.R. Crison, A theoretical basis for a
biopharmaceutic drug classication: the correlation of in vitro drug product
dissolution and in vivo bioavailability, Pharm. Res. 12 (1995) 413420.
[5] L.X. Yu, G.L. Amidon, J.E. Polli, H. Zhao, M.U. Mehta, D.P. Conner, V.P. Shah, L.J.
Lesko, M.L. Chen, V.H.L. Lee, A.S. Hussain, Biopharmaceutics classication system:
the scientic basis for biowaiver extensions, Pharm. Res. 19 (2002) 921925.
[6] C.Y. Wu, L.Z. Benet, Predicting drug disposition via application of BCS: transport/
absorption/ elimination interplay and development of a biopharmaceutics drug
disposition classication system, Pharm. Res. 22 (2005) 1123.
[7] N.A. Kasim, M. Whitehouse, C. Ramachandran, M. Bermejo, H. Lennerns, A.S.
Hussain, H.E. Junginger, S.A. Stavchansky, K.K. Midha, V.P. Shah, G.L. Amidon,
Molecular properties of WHO essential drugs and provisional biopharmaceutical
classication, Mol. Pharmacol. 1 (2004) 8596.
[8] G.C. Williams, P.J. Sinko, Oral absorption of the HIV protease inhibitors: a current
update, Adv. Drug Deliv. Rev. 39 (1999) 211238.
[9] A. Alexaki, Y. Liu, B. Wigdahl, Cellular reservoirs of HIV-1 and their role in viral
persistence, Curr. HIV Educ. Res. 6 (2008) 388400.
[10] L. Varatharajan, S.A. Thomas, The transport of anti-HIV drugs across bloodCNS
interfaces: summary of current knowledge and recommendations for further
research, Antivir. Res. (2009), doi:10.1016/j.antiviral.2008.12.013.
740
741
742
743
744
745
746
747
748
749
750
751
752
753
754
755
756
757
758
759
760
761
762
763
764
765Q8
ARTICLE IN PRESS
RO
OF
[41] R. Lledo-Garcia, A. Nacher, L. Prats-Garcia, V.G. Casabo, M. Merino-Sanjuan,

Bioavailability and pharmacokinetic model for ritonavir in the rat, J. Pharm. Sci.
96 (2007) 633643.
[42] M. Richter, N. Gyemant, J. Molnar, A. Hilgeroth, Comparative effects on intestinal
absorption in situ by P-glycoprotein-modifying HIV protease inhibitors, Pharm.
Res. 21 (2004) 18621866.
[43] L. Tong, T.K. Phan, K.L. Robinson, D. Babusis, R. Strab, S. Bhoopathy, I.J. Hidalgo, G.R.
Rhodes, A.S. Ray, Effects of human immunodeciency virus protease inhibitors on
the intestinal absorption of tenofovir disoproxil fumarate in vitro, Antimicrob.
Agents Chemother. 51 (2007) 34983504.
[44] H.H. Usansky, P. Hu, P.J. Sinko, Differential roles of P-glycoprotein, multidrug
resistance-associated protein 2, and CYP3A on saquinavir oral absorption in
SpragueDawley rats, Drug Metab. Dispos. 36 (2008) 863869.
[45] M.T. Huisman, J.W. Smit, H.R. Wiltshire, R.M.W. Hoetelmans, J.H. Beijnen, A.H.
Schinkel, P-glycoprotein limits oral availability, brain, and fetal penetration of
saquinavir even with high doses of ritonavir, Mol. Pharmacol. 59 (2001) 806813.
[46] J.A. Zastre, G.N.Y. Chan, P.T. Ronaldson, M. Ramaswamy, P.O. Couraud, I.A.
Romero, B. Weksler, M. Bendayan, R. Bendayan, Up-regulation of p-glycoprotein
by HIV protease inhibitors in a human brain microvessel endothelial cell line,
J. Neurosci. Res. 87 (2009) 10231036.
[47] J.W. Polli, J.L. Jarrett, S.D. Studenberg, J.E. Humphreys, S.W. Dennis, K.R. Brouwer,
J.L. Woolley, Role of P-glycoprotein on the CNS disposition of amprenavir
(141W94), an HIV protease inhibitor, Pharm. Res. 16 (1999) 12061212.
[48] M. Johnson, B. Grinsztejn, C. Rodriguez, J. Coco, E. DeJesus, A. Lazzarin, K.
Lichtenstein, A. Rightmire, S. Sankoh, R. Wilber, Atazanavir plus ritonavir or
saquinavir, and lopinavir/ritonavir in patients experiencing multiple virological
failures, AIDS 19 (2005) 685694.
[49] E.S. Perloff, S.X. Duan, P.R. Skolnik, D.J. Greenblatt, L.L. Von Moltke, Atazanavir:
effects on P-glycoprotein transport and CYP3A metabolism in vitro, Drug Metab.
Dispos. 33 (2005) 764770.
[50] E. Ribera, C. Azuaje, R.M. Lopez, M. Diaz, M. Feijoo, L. Pou, M. Crespo, A. Curran, I.
Ocana, A. Pahissa, Atazanavir and lopinavir/ritonavir: pharmacokinetics, safety
and efcacy of a promising double-boosted protease inhibitor regimen, AIDS 20
(2006) 11311139.
[51] B. Young, M.A. Fischl, H.M. Wilson, T.S. Finn, E.H. Jensen, M.J. DiNubile, R.K. Zeldin,
Open-label study of a twice-daily indinavir 800-mg/ritonavir 100-mg regimen in
protease inhibitor-naive HIV-infected adults, J. Acquir. Immune Dec. Syndr. 31
(2002) 478482.
[52] A. Sosnik, D.A. Chiappetta, A.M. Carcaboso, Drug delivery systems in HIV
pharmacotherapy: what has been done and the challenges standing ahead,
J. Control. Release (2009).
[53] V.Y. Alakhov, E.Y. Moskaleva, E.V. Batrakova, A.V. Kabanov, Hypersensitization of
multidrug resistant human ovarian carcinoma cells by pluronic P85 block
copolymer, Bioconjug. Chem. 7 (1996) 209216.
[54] E.V. Batrakova, H.Y. Han, V.Y. Alakhov, D.W. Miller, A.V. Kabanov, Effects of
pluronic block copolymers on drug absorption in Caco-2 cell monolayers, Pharm.
Res. 15 (1998) 850855.
[55] T.J. Spitzenberger, D. Heilman, C. Diekmann, E.V. Batrakova, A.V. Kabanov, H.E.
Gendelman, W.F. Elmquist, Y. Persidsky, Novel delivery system enhances efcacy
of antiretroviral therapy in animal model for HIV-1 encephalitis, J. Cereb. Blood
Flow Metab. 27 (2007) 10331042.
[56] E.V. Batrakova, A.V. Kabanov, Pluronic block copolymers: evolution of drug
delivery concept from inert nanocarriers to biological response modiers,
J. Control. Release 130 (2008) 98106.
[57] Y.C. Kuo, H.H. Chen, Effect of nanoparticulate polybutylcyanoacrylate and
methylmethacrylate-sulfopropylmethacrylate on the permeability of zidovudine
and lamivudine across the in vitro bloodbrain barrier, Int. J. Pharm. 327 (2006)
160169.
[58] Y.C. Kuo, C.Y. Kuo, Electromagnetic interference in the permeability of saquinavir
across the bloodbrain barrier using nanoparticulate carriers, Int. J. Pharm. 351
(2008) 271281.
[59] R. Lobenberg, L. Araujo, H. Von Briesen, E. Rodgers, J. Kreuter, Body distribution of
azidothymidine bound to hexyl-cyanoacrylate nanoparticles after i.v. injection to
rats, J. Control. Release 50 (1998) 2130.
[60] R. Lobenberg, J. Kreuter, Macrophage targeting of azidothymidine: a promising
strategy for AIDS therapy, AIDS Res. Hum. Retrovir. 12 (1996) 17091715.
[61] A. Zensi, D. Begley, C. Pontikis, C. Legros, L. Mihoreanu, S. Wagner, C. Buchel, H.
von Briesen, J. Kreuter, Albumin nanoparticles targeted with Apo E enter the CNS
by transcytosis and are delivered to neurones, J. Control. Release (2009),
doi:10.1016/j.jconrel.2009.03.002.
[62] T. Govender, E. Ojewole, P. Naidoo, I. Mackraj, Polymeric nanoparticles for
enhancing antiretroviral drug therapy, Drug Deliv. 15 (2008) 493501.
[63] E. Ojewole, I. Mackraj, P. Naidoo, T. Govender, Exploring the use of novel drug
delivery systems for antiretroviral drugs, Eur. J. Pharm. Biopharm. 70 (2008)
697710.
[64] V. Schafer, H. Von Briesen, R. Andreesen, A.M. Steffan, C. Royer, S. Troster, J.
Kreuter, H. Rubsamen-Waigmann, Phagocytosis of nanoparticles by human
immunodeciency virus (HIV)-infected macrophages: a possibility for antiviral
drug targeting, Pharm. Res. 9 (1992) 541546.
[65] L.K. Shah, M.M. Amiji, Intracellular delivery of saquinavir in biodegradable
polymeric nanoparticles for HIV/AIDS, Pharm. Res. 23 (2006) 26382645.
[66] M.S. Giri, M. Nebozyhn, A. Raymond, B. Gekonge, A. Hancock, S. Creer, C. Nicols,
M. Yousef, A.S. Foulkes, K. Mounzer, J. Shull, G. Silvestri, J. Kostman, R.G. Collman,
L. Showe, L.J. Montaner, Circulating monocytes in HIV-1-infected viremic
subjects exhibit an antiapoptosis gene signature and virus- and host-mediated
apoptosis resistance, J. Immunol. 182 (2009) 44594470.
CO
RR
EC
TE
DP
[11] D.A. Chiappetta, A. Sosnik, Poly(ethylene oxide)poly(propylene oxide) block

copolymer micelles as drug delivery agents: improved hydrosolubility, stability
and bioavailability of drugs, Eur. J. Pharm. Biopharm. 66 (2007) 303317.
[12] E. Merisko-Liversidge, G.G. Liversidge, E.R. Cooper, Nanosizing: a formulation
approach for poorly-water-soluble compounds, Eur. J. Pharm. Sci. 18 (2003)
113120.
[13] C.C. Lee, J.A. MacKay, J.M.J. Frchet, F.C. Szoka, Designing dendrimers for
biological applications, Nat. Biotechnol. 23 (2005) 15171526.
[14] V.P. Torchilin, Structure and design of polymeric surfactant-based drug delivery
systems, J. Control. Release 73 (2001) 137172.
[15] V.P. Torchilin, Block copolymer micelles as a solution for drug delivery problems,
Expert Opin. Ther. Pat. 15 (2005) 6375.
[16] V.P. Torchilin, Micellar nanocarriers: pharmaceutical perspectives, Pharm. Res.
24 (2007) 116.
[17] V.P. Torchilin, V. Weissig, Polymeric micelles for the delivery of poorly soluble
drugs, ACS Symp. Ser. 752 (2000) 297313.
[18] R.H. Muller, C. Jacobs, O. Kayser, Nanosuspensions as particulate drug formulations in therapy. Rationale for development and what we can expect for the
future, Adv. Drug Deliv. Rev. 47 (2001) 319.
[19] C. Galli, Experimental determination of the diffusion boundary layer width of
micron and submicron particles, Int. J. Pharm. 313 (2006) 114122.
[20] B. Van Eerdenbrugh, L. Froyen, J.A. Martens, N. Blaton, P. Augustijns, M. Brewster,
G. Van den Mooter, Characterization of physico-chemical properties and
pharmaceutical performance of sucrose co-freeze-dried solid nanoparticulate
powders of the anti-HIV agent loviride prepared by media milling, Int. J. Pharm.
338 (2007) 198206.
[21] M.G. Fakes, B.J. Vakkalagadda, F. Qian, S. Desikan, R.B. Gandhi, C. Lai, A. Hsieh, M.
K. Franchini, H. Toale, J. Brown, Enhancement of oral bioavailability of an HIVattachment inhibitor by nanosizing and amorphous formulation approaches, Int.
J. Pharm. 370 (2009) 167174.
[22] Y. Wu, A. Loper, E. Landis, L. Hettrick, L. Novak, K. Lynn, C. Chen, K. Thompson, R.
Higgins, U. Batra, S. Shelukar, G. Kwei, D. Storey, The role of biopharmaceutics in
the development of a clinical nanoparticle formulation of MK-0869: a Beagle dog
model predicts improved bioavailability and diminished food effect on
absorption in human, Int. J. Pharm. 285 (2004) 135146.
[23] E. Merisko-Liversidge, G.G. Liversidge, E.R. Cooper, Nanosizing: a formulation
approach for poorly-water-soluble compounds, Eur. J. Pharm. Sci. 18 (2003)
113120.
[24] P.P. Constantinides, M.V. Chaubal, R. Shorr, Advances in lipid nanodispersions for
parenteral drug delivery and targeting, Adv. Drug Deliv. Rev. 60 (2008) 757767.
[25] H. Dou, J. Morehead, C.J. Destache, J.D. Kingsley, L. Shlyakhtenko, Y. Zhou, M.
Chaubal, J. Werling, J. Kipp, B.E. Rabinow, H.E. Gendelman, Laboratory investigations for the morphologic, pharmacokinetic, and anti-retroviral properties of
indinavir nanoparticles in human monocyte-derived macrophages, Virology 358
(2007) 148158.
[26] L. Baert, G. van 't Klooster, W. Dries, M. Francois, A. Wouters, E. Basstanie, K.
Iterbeke, F. Stappers, P. Stevens, L. Schueller, P. Van Remoortere, G. Kraus, P.
Wigerinck, J. Rosier, Development of a long-acting injectable formulation with
nanoparticles of rilpivirine (TMC278) for HIV treatment, Eur. J. Pharm. Biopharm.
(2009), doi:10.1016/j.ejpb.2009.03.006.
[27] G.G. Liversidge, K.C. Cundy, J.F. Bishop, D.A. Czekai, Surface modied drug
nanoparticles, United States Patent 5, 145, 6841992.
[28] K. Bottomley, Nanotechnology for drug delivery: a validated technology? Drug
Delivery Report Autumn/Winter, 2006, pp. 2021.
[29] A. Martin, P. Bustamante, A.H.C. Chun, Interfacial phenomenon, Physical
Pharmacy, Lippincott Williams & Wilkins, Maryland, 1993, pp. 362392.
[30] M.L. Adams, A. Lavasanifar, G.S. Kwon, Amphiphilic block copolymers for drug
delivery, J. Pharm. Sci. 92 (2003) 13431355.
[31] M.L. Adams, A. Lavasanifar, G.S. Kwon, Erratum: amphiphilic block copolymers
for drug delivery (Journal of Pharmaceutical Research (2003) 92, 7 (13431355)
doi:10.1002/jps.10397), J. Pharm. Sci. 94 (2005) 1160.
[32] L. Bromberg, Polymeric micelles in oral chemotherapy, J. Control. Release 128
(2008) 99112.
[33] Y. Nagasaki, K. Yasugi, Y. Yamamoto, A. Harada, K. Kataoka, Sugar-installed block
copolymer micelles: their preparation and specic interaction with lectin
molecules, Biomacromolecules 2 (2001) 10671070.
[34] C. Allen, D. Maysinger, A. Eisenberg, Nano-engineering block copolymer
aggregates for drug delivery, Colloids Surf., B. Biointerfaces 16 (1999) 327.
[35] H.M. Aliabadi, M. Shahin, D.R. Brocks, A. Lavasanifar, Disposition of drugs in block
copolymer micelle delivery systems: from discovery to recovery, Clin. Pharmacokinet. 47 (2008) 619634.
[36] N. Shaik, G. Pan, W.F. Elmquist, Interactions of pluronic block copolymers on P-gp
efux activity: experience with HIV-1 protease inhibitors, J. Pharm. Sci. 97 (2008)
54215433.
[37] E.V. Batrakova, S. Li, D.W. Miller, A.V. Kabanov, Pluronic P85 increases
permeability of a broad spectrum of drugs in polarized BBMEC and Caco-2 cell
monolayers, Pharm. Res. 16 (1999) 13661372.
[38] E.V. Batrakova, D.W. Miller, S. Li, V.Y. Alakhov, A.V. Kabanov, W.F. Elmquist,
Pluronic P85 enhances the delivery of digoxin to the brain: in vitro and in vivo
studies, J. Pharmacol. Exp. Ther. 296 (2001) 551557.
[39] D.W. Miller, E.V. Batrakova, A.V. Kabanov, Inhibition of multidrug resistanceassociated protein (MRP) functional activity with pluronic block copolymers,
Pharm. Res. 16 (1999) 396401.
[40] R.B. Kim, M.F. Fromm, C. Wandel, B. Leake, A.J.J. Wood, D.M. Roden, G.R.
Wilkinson, The drug transporter P-glycoprotein limits oral absorption and brain
entry of HIV-1 protease inhibitors, J. Clin. Invest. 101 (1998) 289294.
UN
766
767
768
769
770
771
772
773
774
775
776
777
778
779
780
781
782
783
784
785
786
787
788
789
790
791
792
793
794
795
796
797
798
799
800
801
802
803
804
805
806
807
808
809
810
811
812
813
Q9 814
815
816
817
818
819
820
821
822
823
824
825
826
827
828
829
830
831
832
833
834
835
836
837
838
839
840
841
842
843
844
845
846
847
848
849
850
851
11
852
853
854
855
856
857
858
859
860
861
862
863
864
865
866
867
868
869
870
871
872
873
874
875
876
877
878
879
880
881
882
883
884
885
886
887
888
889
890
891
892Q10
893
894
895
896
897
898
899
900
901
902
903
904
905
906
907
908
909
910
911
912
913
914
915
916
917
918
919
920Q11
921
922
923
924
925
926
927
928
929
930
931
932
933
934
935
936
937
ARTICLE IN PRESS
[92]
[93]
[94]
[95]
[96]
[97]
[98]
[99]
[100]
[101]
[102]
OF
[91]
RO
[90]
inuence of size, charge and uorescent labeling, Pharm. Res. 23 (2006) 999
28182826.
1000
D.S. Pisal, V.K. Yellepeddi, A. Kumar, R.S. Kaushik, M.B. Hildreth, X. Guan, S. 1001
Palakurthi, Permeability of surface-modied polyamidoamine (PAMAM) den- 1002
drimers across Caco-2 cell monolayers, Int. J. Pharm. 350 (2008) 113121.
1003
K.M. Kitchens, M.E.H. El-Sayed, H. Ghandehari, Transepithelial and endothelial 1004
transport of poly (amidoamine) dendrimers, Adv. Drug Deliv. Rev. 57 (2005) 1005
21632176.
1006
A. D'Emanuele, R. Jevprasesphant, J. Penny, D. Attwood, The use of a dendrimer- 1007
propranolol prodrug to bypass efux transporters and enhance oral bioavail- 1008
ability, J. Control. Release 95 (2004) 447453.
1009
R. Wiwattanapatapee, B. Carreno-Gomez, N. Malik, R. Duncan, Anionic PAMAM 1010
dendrimers rapidly cross adult rat intestine in vitro: a potential oral delivery 1011
system? Pharm. Res. 17 (2000) 991998.
1012
K.L. Aillon, Y. Xie, N. El-Gendy, C.J. Berkland, M.L. Forrest, Effects of nanomaterial 1013
physicochemical properties on in vivo toxicity, Adv. Drug Deliv. Rev. 61 (2009) 1014
457466.
1015
R. Jevprasesphant, J. Penny, D. Attwood, N.B. McKeown, A. D'Emanuele, 1016
Engineering of dendrimer surfaces to enhance transepithelial transport and 1017
reduce cytotoxicity, Pharm. Res. 20 (2003) 15431550.
1018
R.B. Kolhatkar, K.M. Kitchens, P.W. Swaan, H. Ghandehari, Surface acetylation of 1019
polyamidoamine (PAMAM) dendrimers decreases cytotoxicity while maintain- 1020
ing membrane permeability, Bioconjug. Chem. 18 (2007) 20542060.
1021
M.F. Neerman, W. Zhang, A.R. Parrish, E.E. Simanek, In vitro and in vivo 1022
evaluation of a melamine dendrimer as a vehicle for drug delivery, Int. J. Pharm. 1023
281 (2004) 129132.
1024
H.T. Chen, M.F. Neerman, A.R. Parrish, E.E. Simanek, Cytotoxicity, hemolysis, and 1025
acute in vivo toxicity of dendrimers based on melamine, candidate vehicles for 1026
drug delivery, J. Am. Chem. Soc. 126 (2004) 1004410048.
1027
J. Whelan, First clinical data on RNAi, Drug Discov. Today 10 (2005) 10141015. 1028
M. Keller, Nanomedicinal delivery approaches for therapeutic siRNA, Int. J. 1029
Pharm. (2009).
1030
Q12
J. Capodici, K. Karik, D. Weissman, Inhibition of HIV-1 infection by small 1031
interfering RNA-mediated RNA interference, J. Immunol. 169 (2002) 51965201. 1032
W.Y. Hu, C.P. Myers, J.M. Kilzer, S.L. Pfaff, F.D. Bushman, Inhibition of retroviral 1033
pathogenesis by RNA interference, Curr. Biol. 12 (2002) 13011311.
1034
J.M. Jacque, K. Triques, M. Stevenson, Modulation of HIV-1 replication by RNA 1035
interference, Nature 418 (2002) 435438.
1036
S. Mao, M. Neu, O. Germershaus, O. Merkel, J. Sitterberg, U. Bakowsky, T. Kissel, 1037
Inuence of polyethylene glycol chain length on the physicochemical and 1038
biological properties of poly(ethylene imine)-graft-poly(ethylene glycol) block 1039
copolymer/SiRNA polyplexes, Bioconjug. Chem. 17 (2006) 12091218.
1040
N. Weber, P. Ortega, M.I. Clemente, D. Shcharbin, M. Bryszewska, F.J. de la Mata, R. 1041
Gmez, M.A. Muoz-Fernndez, Characterization of carbosilane dendrimers as 1042
effective carriers of siRNA to HIV-infected lymphocytes, J. Control. Release 132 1043
(2008) 5564.
1044
T. Dutta, M. Garg, N.K. Jain, Targeting of efavirenz loaded tuftsin conjugated poly 1045
(propyleneimine) dendrimers to HIV infected macrophages in vitro, Eur. J. 1046
Pharm. Sci. 34 (2008) 181189.
1047
T. Dutta, N.K. Jain, Targeting potential and anti-HIV activity of lamivudine loaded 1048
mannosylated poly (propyleneimine) dendrimer, Biochim. Biophys. Acta, Gen. 1049
Subj. 1770 (2007) 681686.
1050
Z. Cui, C.H. Hsu, R.J. Mumper, Physical characterization and macrophage cell 1051
uptake of mannan-coated nanoparticles, Drug Dev. Ind. Pharm. 29 (2003) 1052
689700.
1053
L. Margolis, R. Shattock, Selective transmission of CCR5-utilizing HIV-1: the 1054
gatekeeper problem resolved? Nat. Rev. Microbiol. 4 (2006) 312317.
1055
P.J. Klasse, R. Shattock, J.P. Moore, Antiretroviral drug-based microbicides to 1056
prevent HIV-1 sexual transmission, Annu. Rev. Med. 59 (2008) 455471.
1057
[103]
[104]
[105]
TE
[67] J.M. Orenstein, The macrophage in HIV infection, Immunobiology 204 (2001)
598602.
[68] R.A. Weiss, How does HIV cause AIDS? Science 260 (1993) 12731279.
[69] J.M. Orenstein, C. Fox, S.M. Wahl, Macrophages as a source of HIV during
opportunistic infections, Science 276 (1997) 18571860.
[70] F. Chellat, Y. Merhi, A. Moreau, L. Yahia, Therapeutic potential of nanoparticulate
systems for macrophage targeting, Biomaterials 26 (2005) 72607275.
[71] D. Leu, B. Manthey, J. Kreuter, Distribution and elimination of coated polymethyl
[2-14C]methacrylate nanoparticles after intravenous injection in rats, J. Pharm.
Sci. 73 (1984) 14331437.
[72] F. Pierige, S. Serani, L. Rossi, M. Magnani, Cell-based drug delivery, Adv. Drug
Deliv. Rev. 60 (2008) 286295.
[73] S.M. Moghimi, A.C. Hunter, J.C. Murray, Long-circulating and target-specic
nanoparticles: theory to practice, Pharmacol. Rev. 53 (2001) 283318.
[74] G. Storm, S.O. Belliot, T. Daemen, D.D. Lasic, Surface modication of nanoparticles
to oppose uptake by the mononuclear phagocyte system, Adv. Drug Deliv. Rev. 17
(1995) 3148.
[75] D.E. Owens III, N.A. Peppas, Opsonization, biodistribution, and pharmacokinetics
of polymeric nanoparticles, Int. J. Pharm. 307 (2006) 93102.
[76] A. Vonarbourg, C. Passirani, P. Saulnier, J.P. Benoit, Parameters inuencing the
stealthiness of colloidal drug delivery systems, Biomaterials 27 (2006)
43564373.
[77] A.S. Ham, M.R. Cost, A.B. Sassi, C.S. Dezzutti, L.C. Rohan, Targeted delivery of PSCRANTES for HIV-1 prevention using biodegradable nanoparticles, Pharm. Res. 26
(2009) 502511.
[78] K.A. Woodrow, Y. Cu, C.J. Booth, J.K. Saucier-Sawyer, M.J. Wood, W. Mark
Saltzman, Intravaginal gene silencing using biodegradable polymer nanoparticles densely loaded with small-interfering RNA, Nat. Mater. 8 (2009) 526533.
[79] M.P. Desai, V. Labhasetwar, G.L. Amidon, R.J. Levy, Gastrointestinal uptake of
biodegradable microparticles: effect of particle size, Pharm. Res. 13 (1996)
18381845.
[80] S.K. Lai, D.E. O'Hanlon, S. Harrold, S.T. Man, Y.Y. Wang, R. Cone, J. Hanes, Rapid
transport of large polymeric nanoparticles in fresh undiluted human mucus,
Proc. Natl. Acad. Sci. U. S. A. 104 (2007) 14821487.
[81] Y. Cu, W.M. Saltzman, Controlled surface modication with poly(ethylene)glycol
enhances diffusion of PLGA nanoparticles in human cervical mucus, Mol.
Pharmacol. 6 (2009) 173181.
[82] D.A. Chiappetta, A.M. Carcaboso, C. Bregni, M. Rubio, G. Bramuglia, A. Sosnik,
Indinavir-loaded pH-sensitive microparticles for taste masking: toward extemporaneous pediatric anti-HIV/AIDS liquid formulations with improved patient
compliance, AAPS PharmSciTech 10 (2009) 16.
[83] C. Villalonga-Barber, M. Micha-Screttas, B.R. Steele, A. Georgopolous, C.
Demetzos, Dendrimers as biopharmaceuticals: synthesis and properties, Curr.
Top. Med. Chem. 8 (2008) 12941309.
[84] M. El-Sayed, M. Ginski, C. Rhodes, H. Ghandehari, Transepithelial transport of
poly(amidoamine) dendrimers across Caco-2 cell monolayers, J. Control. Release
81 (2002) 355365.
[85] M. El-Sayed, M. Ginski, C.A. Rhodes, H. Ghandehari, Inuence of surface
chemistry of poly(amidoamine) dendrimers on Caco-2 cell monolayers, J. Bioact.
Compat. Polym. 18 (2003) 722.
[86] M. El-Sayed, C.A. Rhodes, M. Ginski, H. Ghandehari, Transport mechanism(s) of
poly (amidoamine) dendrimers across Caco-2 cell monolayers, Int. J. Pharm. 265
(2003) 151157.
[87] R. Jevprasesphant, J. Penny, R. Jalal, D. Attwood, N.B. McKeown, A. D'Emanuele,
The inuence of surface modication on the cytotoxicity of PAMAM dendrimers,
Int. J. Pharm. 252 (2003) 263266.
[88] K.M. Kitchens, A.B. Foraker, R.B. Kolhatkar, P.W. Swaan, H. Ghandehari,
Endocytosis and interaction of poly (amidoamine) dendrimers with Caco-2
cells, Pharm. Res. 24 (2007) 21382145.
[89] K.M. Kitchens, R.B. Kolhatkar, P.W. Swaan, N.D. Eddington, H. Ghandehari,
Transport of poly(amidoamine) dendrimers across Caco-2 cell monolayers:
[106]
RR
EC
[107]
[108]
[109]
CO
[110]
UN
938
939
940
941
942
943
944
945
946
947
948
949
950
951
952
953
954
955
956
957
958
959
960
961
962
963
964
965
966
967
968
969
970
971
972
973
974
975
976
977
978
979
980
981
982
983
984
985
986
987
988
989
990
991
992
993
994
995
996
997
998
1058
DP
12

Pure Drug and Polymer Based Nanotechnologies For The Improved Solubility, Stability

Uploaded by

Document Information

Original Title

Copyright

Available Formats

Share this document

Share or Embed Document

Sharing Options

Did you find this document useful?

Is this content inappropriate?

Copyright:

Available Formats

Pure Drug and Polymer Based Nanotechnologies For The Improved Solubility, Stability

Uploaded by

Copyright:

Available Formats

ARTICLE IN PRESS

Advanced Drug Delivery Reviews xxx (2009) xxxxxx

Contents lists available at ScienceDirect

Advanced Drug Delivery Reviews

Puneet Sharma, Sanjay Garg

School of Pharmacy, The University of Auckland, Auckland, New Zealand

Human immunodeciency virus (HIV) related acquired immune

treatment or HAART, is considered as one of the most signicant

2. Pure drug nanoparticles

In a pure drug nanoparticle formulation, submicron size particles

the extent of the intestinal absorption (parent drug plus metabolites)

P. Sharma, S. Garg / Advanced Drug Delivery Reviews xxx (2009) xxxxxx

Entry and fusion

b300 nm. In the process, the drug suspension containing macro or

nanosized loviride formulation which was freeze-dried with sucrose

Papp AP to BLb (cm/s)

Aq: puried water. NR: not reported.

3. Polymer based nanotechnologies

3.1. Polymeric micelles

Polymeric micelles are nanostructures of b100 nm diameter that

rst-pass metabolism and results in dose reduction which is highly

nanoparticles reduces the bioavailability variability brought by the

P. Sharma, S. Garg / Advanced Drug Delivery Reviews xxx (2009) xxxxxx

3.2. Polymeric nanoparticles

Polymeric nanoparticles (NPs) are b1000 nm in size and are

0.001% and 0.01% showed signicant reduction in HIV replication.

micelles is relatively high [16,32]. Polymeric micelles consist of a

Fig. 2. Pluronic block copolymer P85 inhibits HIV-1 replication in monocyte-derived

[62,63]. In most cases, polymeric nanoparticles have been used to

nanoparticles. In addition, transportation of apolipoprotein bound

P. Sharma, S. Garg / Advanced Drug Delivery Reviews xxx (2009) xxxxxx

A dendrimer is a polymeric nanostructure (b100 nm) built around

concentrations. Taste masking studies were conducted on healthy

P. Sharma, S. Garg / Advanced Drug Delivery Reviews xxx (2009) xxxxxx

P. Sharma, S. Garg / Advanced Drug Delivery Reviews xxx (2009) xxxxxx

4. Conclusion and future directions

The growing number of HIV infected individuals and related

impaired biodistribution in HIV reservoirs, emergence of drug

vivo studies. With the establishment of the proof of concept, there is a

Fig. 8. Transfection and toxicity proles of dendriplexes on HIV infected T cells

Fig. 9. Sequence specic inhibition of HIV in PBMC after 24 h incubation with

[41] R. Lledo-Garcia, A. Nacher, L. Prats-Garcia, V.G. Casabo, M. Merino-Sanjuan,

[11] D.A. Chiappetta, A. Sosnik, Poly(ethylene oxide)poly(propylene oxide) block

P. Sharma, S. Garg / Advanced Drug Delivery Reviews xxx (2009) xxxxxx

You might also like