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CURRENT PROSPECTS OF NANO-DESIGNS IN GENE DELIVERY- AIMING NEW HIGH FOR EFFICIENT AND TARGETED GENE THERAPY

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The twentieth century is considered for the technological revolution in different fields such as Industry, Research and Medicine. Scientific inventions have improved Research and Industrial output while medicine one step ahead. Numerous inventions have made a revolution in the management of life threatening diseases which were incurable decades ago. Due to these revelations, health care system is growing exponentially and especially, Gene Therapy is known as one of the most advanced approaches for the treatment of diseases associated with abnormal functioning of the genome. Gene therapy offers management of diseases/disorders through manipulation at genetic level either by replacement of abnormal gene(s) and /or repairs. Gene therapy essentially requires targeted and efficient gene delivery to tissue or cell. There are numerous methods available to carry out gene delivery either In-Vivo or Ex-Vivo for particular diseases. Both viral and non-viral vehicles used for delivery of exogenous genes have shown tremendous benefits in numerous clinical trials carried out over last few decades. Both the options, viral and non-viral tools, for gene delivery with remarkable significance are often linked with numerous complications. To surpass these complications, novel tools like Nano-constructs designed for nano-materials are in practice and have shown promising results in delivering candidate drugs and biomolecules. Here in this review, we have summarized the potential of new generation delivery vehicles and their advantages over viral and other available non-viral vehicles. Also, current information with respect to the design and functioning of Nano-constructs implemented in clinical study for management of many diseases is provided.
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Mahendra Kumar Verma. et al. / International Journal of Biopharmaceutics. 2013; 4(3): 145-165.
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CURRENT PROSPECTS OF NANO-DESIGNS IN GENE DELIVERY-
AIMING NEW HIGH FOR EFFICIENT AND TARGETED GENE
THERAPY
Mahendra Kumar Verma
1
, Yogendra Kumar Verma
2
, Kota Sobha
*3
,
Subrato Kumar Dey
4
1&*3
Department of Biotechnology, R.V.R. & J.C. College of Engineering (A), Guntur 522 019, Andhra Pradesh, India.
2
Department of Neurobiology, All India Institute of Medical Sciences, New Delhi 110 029, India.
4
School of Biological Sciences, West Bengal University of Technology, Kolkata 700 064, West Bengal, India.
ABSTRACT
The twentieth century is considered for the technological revolution in different fields such as Industry, Research
and Medicine. Scientific inventions have improved Research and Industrial output while medicine one step ahead.
Numerous inventions have made a revolution in the management of life threatening diseases which were incurable decades
ago. Due to these revelations, health care system is growing exponentially and especially, Gene Therapy is known as one of
the most advanced approaches for the treatment of diseases associated with abnormal functioning of the genome. Gene
therapy offers management of diseases/disorders through manipulation at genetic level either by replacement of abnormal
gene(s) and /or repairs. Gene therapy essentially requires targeted and efficient gene delivery to tissue or cell. There are
numerous methods available to carry out gene delivery either In-Vivo or Ex-Vivo for particular diseases. Both viral and non-
viral vehicles used for delivery of exogenous genes have shown tremendous benefits in numerous clinical trials carried out
over last few decades. Both the options, viral and non-viral tools, for gene delivery with remarkable significance are often
linked with numerous complications. To surpass these complications, novel tools like Nano-constructs designed for nano-
materials are in practice and have shown promising results in delivering candidate drugs and biomolecules. Here in this
review, we have summarized the potential of new generation delivery vehicles and their advantages over viral and other
available non-viral vehicles. Also, current information with respect to the design and functioning of Nano-constructs
implemented in clinical study for management of many diseases is provided.
Key words: Gene Delivery, Non-viral and Viral vehicles, Transformation efficiency, Nanomaterial, Nano-constructs.
INTRODUCTION
Gene Therapy, a multidisciplinary approach
offers treatment for numerous diseases and disorders that
arise due to failure of gene function. Gene therapy, a
frontier medicine and growing exponentially, provides
Corresponding Author
Dr. Kota Sobha
Email: sobhak66@gmail.com
treatment for genetic disorders (Cavazzana-Calvo, 2000).
In the year 1972, first attempt had been made in the field
of medicine beyond conventional methods. It was a
breakthrough in the conventional healthcare system after
a trial of gene therapy for the management of Severe
Combined Immunodeficiency Syndrome (SCID)
(Hacein-Bey-Abina et al., 2002; 2003). Based on the
clinical output of SCID in 1972, numerous clinical trials
have been made in subsequent years for treatment of
different diseases and metabolic disorders (Roman
Gardlík et al., 2005). Advancement of technology
IJB
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especially in the areas of molecular biology, recombinant
DNA technology and proteomics led Gene Therapy to be
a refined, more efficient and popular part of current
healthcare systems (Fioretti et al., 2013; Pavel Simara et
al., 2013).
In the present scenario with rapid technological
revolution, health care system has grown tremendously
but still there are a number of diseases which are still
incurable.
The major challenge with the diseases like
genetic disorders and metabolic disorders is to unravel
the cause(s) at molecular level especially at genetic level
(Maria et al., 2013; Rick et al, 2013). Genetic defects and
metabolic disorders cannot be cured by conventional
approach of medicine but only symptomatic relief could
be achieved. Hence the importance of alternate
medication such as Gene Therapy is pertinent to find out
the prime cause of disease and its management (Micaela
Johanna Glat and Daniel Offen, 2013; Blagbrough and
Zara, 2009; Guo and Huang, 2012). Diabetes mellitus has
created a devastating condition in every age group and
possible treatment available is the continuous use of
       
prime choice can be transformed into permanent
treatment via replacement of nonfunctional part of insulin
gene by Gene Therapy (Cummings, 2013; Jasmin
Lebastchi 2013).
PREREQUISITES FOR GENE THERAPY
It became more significant to find out the causes
of diseases / disorders that are often associated with a
higher rate of mortality. Research made us believe that
the flow of genetic information in a proper way is
essential for a healthy living system which otherwise gets
disturbed either by an inborn error or any invading
element (Elena and Mary, 2013; James et al., 2013). In
both the cases, information coded in the genome cannot
transform precisely leading to diseased state (Shyamal,
2013). Gene Therapy works on the principle of
refinement of the genetic construct responsible for
disease or metabolic disorders. The Therapy works two
ways: one is repair of the defective part of the genome
and the other, replacement with fresh functional copy
(Chen-Hsien Su, Deniz Erol, 2013; Basarkar and Singh,
2007). We can achieve gene delivery either Ex-Vivo or
In-Vivo based on the disease under consideration and the
method opted for gene delivery (Atkinson and Chalmers,
2010; Bushman, 2007). Among these approaches,
replacements of the defective or malfunctioning gene(s)
with a fresh copy is more fissile rather than to repair
individual bases in a pool of genome (Sakurai et al.,
2003; Edelstein et al., 2007; Boudreau and Davidson,
2012). The efficiency of Gene therapy depends on
successful delivery of candidate gene to specific and
correct locus in the genome. While carrying out gene
therapy at the clinical level, it becomes more important to
regulate the transformation of fresh copy in a required
copy number without altering the rest of the genome
(Jacobs et al., 2008). Gene transformation efficiency and
its expression are other parameters which define the
efficiency of gene therapy.
CANDIDATE DISEASES FOR GENE THERAPY
The emergence of novel diseases and limitations
of conventional healthcare system signifies Gene therapy
that offers novel options for management of diseases
such as Cystic Fibrosis, SCID, Hemophilia, Diabetes
mellitus, Muscular Dystrophy, Multiple Sclerosis, Sickle
Cell Anemia, Cancer and AIDS that are quite difficult to
cure with conventional medicine (Dai, 2012; Kostense ,
2004; Shi and Pardridge, 2000). Gene therapy is a viable
option for all those diseases/disorders that involve
malfunctioning of a single or multiple genes directly or
indirectly. In order to perform gene therapy for listed
diseases, one needs a carrier molecule that can carry a
fresh copy of the gene and transplant at the appropriate
position in the genome (Guliyeva, 2006; Rao and Gopal,
2006). There are numerous methods available for
targeted gene delivery and all of them are basically
classified into two categories: one viral vehicle and
another, non-viral or physical methods (Liu et al., 1997;
Cichon et al., 2001).
VIRAL VEHICLES
Success of gene therapy depends on efficient
and specific delivery of candidate gene(s) to targeted
tissue or cell. Among the many inventions most
significant breakthrough was the use of viruses, the
obligate intracellular parasites, as tools to carry gene(s)
into host cells after refinement (Kostense et al., 2004;
Cotter et al., 2005). The criterion for choosing a virus as
delivery vehicle is its innate property to infect cells in
higher proportions. An added advantage of viruses as
delivery vehicles is their selectivity towards their host
system (Nayak and Herzog, 2010; Li et al., 2003). Thus
high gene transfer efficiency and selective gene delivery
made virus based vehicles more popular. Both DNA (Ex:
adenoviruses) and RNA viruses (Ex: retrovirus) were
refined at the genetic level to transform into delivery
vehicles (Bushman, 2007). Additionally, a few more
classes of viruses such as Lentivirus, Herpes Simplex
Virus (HSV) and Adeno-Associated Viruses (AAD) also
have shown their advantages in gene delivery. Numerous
virus hybrid systems have also been designed to improve
efficiency of gene delivery and success of gene therapy
(Carlson, 2008; Sliva and Schnierle, 2010).
Although viruses have significant advantages as
gene delivery vehicles, they are often associated with
many life threatening complications (Shyam, 2008;
Bouard, 2009). Since we are dealing with the most
pathogenic elements (Viruses) as treatment strategy for
diseases, the risk of vehicular virus causing disease will
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sustain (Clare et al., 2003; Latchman, 2001). In addition,
viruses and other microorganisms exhibit innate ability to
uptake exogenous genes from the environment through
Horizontal Gene Transfer mechanism (HGT) without
passing into the next generations. Probably HGT is the
prime cause of post gene therapy complications/failures
due to intake of virulence genes (Thomas, 2001).
Immuno-complications are often frequent with viral
based gene delivery reflecting another major drawback
with the viral vehicles (Thomas et al., 2000; Wang,
2001). Further, while viral based vehicles can effectively
transfer a limited size of nucleic acid, many diseases like
SCID need multiple gene transfer at the same time which
becomes a constraint and hence the failure of viral
vehicles.
NON-VIRAL VEHICLES
With frequent complications offered by viral
vehicles, there was an immense need for novel vehicles
with improved efficiency and minimum complications
(Alan et al., 2003). Numerous methods and tools have
been designed to fulfill the requirements for a vehicle to
        
non-viral or physical methods have shown promising
result in clinical implications and commercial use (Xu et
al., 2007). Among the numerous tools based on non-viral
option for gene delivery, the notable ones are Gene Gun,
Direct Gene transfer, Liposomes and Immuno-liposomes
that significantly established their role in the new era of
medicine (Picquet et al., 2005; Ewert et al., 2006). The
most important feature of non-viral based tools is their
safety and ability of carrying unlimited size of nucleic
acid. These methods equipped novel medicine, Gene
Therapy, to deliver multiple genes simultaneously and
safely (Yang et al., 2007; Yoshida et al., (2000). Though
non-viral based gene delivery is considered safe and
effective, it also has its own drawbacks like non-
specificity (McCaffrey et al., 2002; Oupicky et al., 2002)
and immunogenicity effecting clinical complications (Lin
et al. , 2000; Prata et al., 2008). In fact, options like
Gene Gun, direct gene delivery and transformation are
quite good at the laboratory but not clinically and hence
cannot be used in In-Vivo gene therapy (Park and Healy,
2004).
CHALLENGES TO CONVENTIONAL DELIVERY
SYSTEM
Targeted delivery of exogenous gene(s) in
eukaryotic system is much more difficult than
prokaryotic system due to multiple biological boundary
and cellular trafficking (Mohammed et al., 2009; Walther
and Stein, 2000). Success of Gene Therapy depends on
efficient and targeted gene delivery crossing the
biological boundaries and expression of the candidate
gene subsequently. Existing methods for gene delivery
both viral and non-viral are efficient but with notable
post therapy complications (Nguyen et al., 2007; Chen et
al., 2003). Especially viral vehicles often lead to
immunological complications primarily while non-viral
tools often get failed to cross various biological
boundaries and cellular trafficking (Sakai et al., 2005).
An ideal delivery vehicle either viral or non-viral must
possess the enlisted characteristics for successful Gene
Therapy.
The desired features are:
1. Non-toxic, non-pathogenic and non-immunogenic.
2. Must carry large size of exogenous nucleic acid.
3. Ease of administration and high serum stability.
4. Ease in DNA packaging/ Construct Design.
5. High nuclear transport and transformation efficiency.
6. Possess higher specificity and internalization.
7. Must be inexpensive and escape endolysosomes.
8. Ease of Design/ Modification
9. Wide spectrum; must deliver genes to dividing and
non-dividing cells.
10. Must be economical.
It is much difficult to design and maintain
characteristics listed above in a single vehicle for gene
delivery. Both viral and non-viral vehicles often fail to
retain these essential features. Especially viral vehicles
exhibit dynamic genome creating difficulties to
manipulate and lead to high level Immune-complications
(Marti et al., 2004; Yiqi Seow and Matthew, 2009). More
often the stability problem has been a major challenge
with non-viral methods with low specificity and systemic
toxicity. Serum instability and low perfusion rate are
other challenges noticed with non-viral tools for gene
delivery (Oliver et al., 2007). The drawbacks exhibited
by existing vehicles need to be addressed by further
refinement and/or design of new tools for efficient gene
delivery. At this time, Nano-constructs have emerged as
most reliable options for gene therapy (Aneed, 2004).
NANOTECHNOLOGY IN GENE DELIVERY
Nanotechnology offers novel designs by fine
manipulation of materials at Nano-scale very close to
atomic and molecular level (Jeschke and Klein, 2004).
These novel designs offer enormous applications in
different areas such as Material Science, Biological
Science and Research (Ogris and Wagner, 2002). This
technology is ideal to reveal the etiology of molecules
that cross varied biological barriers and track their fate at
subcellular compartments (Vighi, 2010; Ng and Pun,
2008). In the context of Gene Therapy, nanotechnology is
desperately needed as it provides gene delivery beyond
biological boundaries. Programmed uptake of exogenous
nucleic acid (DNA) in a eukaryotic cell needs that
vehicles cross the plasma membrane, subcellular traffic
and nuclear membrane (Viola et al., 2010; Muro et al.,
2006). Nanoscale materials such as nanoparticles,
nanotubes and nanofibers offer higher efficiency of gene
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delivery with minimal loss and post therapy
complications in clinical subjects (Choi et al., 2008;
Dincer et al., 2005).
AVAILABLE NANO-CONSTRUCTS FOR GENE
DELIVERY
From the time of the first designed nanoconstruct
liposome late 1960s till date, numerous Nano-scale
devices have been designed and used clinically as part of
advanced medicine (Ruozi et al., 2003). Nano-constructs
used for gene/drug delivery and grouped as non-viral/
physical/mechanical methods include Liposomes,
Immune-liposomes, Nanopores, Fullerenes, Nanotubes,
Quantum dots, Nanoshells, Nanobubbles, Dendrimers,
Magnetic nanoparticles and Nanosomes. Many of these
have been explored and established as ideal carriers for
drugs while much more optimization is needed for
identification of suitable nanomaterials for gene delivery.
Materials like liposomes, immuno-liposomes, nanotubes,
nanopores and quantum dots have been approved by the
Federal Drug Administration (FDA) for drug and gene
delivery (Mykhaylyk, 2007).
LIPOSOMES: BEGINNING OF NANO-MEDICINE
With the advancement of technology, several
classes of nanomaterials have been designed and used for
medical applications. For example, in cancer
management, chemotherapy and delivery of anticancer
drugs were carried out in the late 1960s using liposomes
(Lasic, 1997; Ito et al., 2004). Liposome was the first
attempt in the late 1960s which was designed and
implemented for drug delivery at nanoscale. Liposomes
are nanoscale spherical devises made of phospholipids
which are amphipathic and arranged into a sphere in 3D
with a hollow central cavity providing enough space for
loading of candidate drugs (Gao and Hui (2001). The
liposome can vary in size and chemical behavior based
on the design/orientation of phospholipids as shown in
Figure 1. Liposome based drug delivery system got
popular in a small span of time due to its advantages over
other available methods (Amarnath and Uma, 1997).
Simultaneously, the stability remains a major
challenge with liposomes as they easily release drugs and
get removed by macrophages especially in liver (Gregory
Gregoriadis, 1995). Many attempts have been made to
improve half-life with the incorporation of chemicals
such as cholesterol, polyvinylpyrollidone,
polyacrylamide lipids and destroy phosphatidylcholine
for thermal stability (Toshinori and Junzo, 1992; Seville,
2002). Both cationic and anionic liposomes are
extensively used for grafting DNA cargoes into cells
(Narang et al., 2005). Liposome based gene delivery is
ubiquitously employed in the management of many
diseases such as cardiovascular disorders, renal
glomerulosclerosis and cancer (Hoyer and Neundorf,
2012). Viral origin fusigenic liposomes were used in in
vitro studies in delivering genes to cardiac, cerebral and
kidney tissues (Zou et al., 2000; Schmitz et al., 2004).
Liposomes offer excellent opportunity to deliver
genes to superficial organs like skin and deeper tissues
like brain as well with enormous potential. Moreover
liposomes in Free State, conjugated with biomolecules
and then lyophilized improve the versatility of therapy
(Li and Huang, 2004; Li et al., 2000). Cationic liposome
tagged with proteins effecting wide spectrum gene
delivery in dried powder form is ideal for skin and
intratracheal disorders (Almofti, 2003). Further, charged
and variant liposomes with the ability of gene delivery to
both dividing and non-dividing cells define their potential
for delivery of gene to neurological and tumor tissues
(Ueno et al., 2002). Integrin mediated liposomes provide
a unique platform for the gene delivery to bone marrow
with enhanced infusion and transfer efficiency (Uddin
and Islam (2006); Uduehi et al., 2003). Low molecular
weight Chitosan based liposomes have shown their
potential in deeper tissues as they could go beyond
biological barriers (Dennis et al., 2003; Lee et al., 2001).
IMMUNO-LIPOSOMES: REFINED VARIANT OF
LIPOSOMES
Refined liposomes that deliver drug and nucleic
acid with specificity are termed immune-liposomes
(Roberta et al., 2012). Design and synthesis of immune-
liposome requires profiling of surface proteins of the
targeted tissue and raise of antibodies against them.
These antibodies hybridized with liposome let specific
delivery of candidate drug and nucleic acid as shown in
figure 2. In the last decade immune-liposomes based drug
delivery was used in the management of cancer and other
diseases and the success of it is largely attributed to its
efficient targeted delivery based on protein-protein
interactions (John et al., 2013). Further, poly-epitopic
and polyclonal antibodies improved specific delivery of
candidate genes and drugs to targeted tissue or cell
(Blumling et al., 2012). With optimized properties of
immuno-liposomes, clinical popularity is increasing
exponentially. A novel variant of immuno-liposomes is in
clinical use for disorders like SCID, Diabetes mellitus,
multiple sclerosis and cystic fibrosis (Balicki et al., 2002;
Puja and Theresa, 2004; Khawa et al., 2001; Makiya
and Huang, 2004; Srinivas et al., 2008; Song et al., 2000;
Purnima et al., 2002; Manuela and Maya, 2007; Maaike
et al., 2005).
With the enhanced potential, often
immunoliposomes are grouped under newer generation
rather than the first generation delivery systems.
Abnormal expression of HER2 oncogenes is a major
cause of breast cancer with high mortality worldwide.
Recently, anti-p185HER2 tagged immunoliposomes with
a tremendous binding affinity with tumor cell in-vivo are
implemented in cancer therapy (John et al., 1996; Dmitri
et al., 2006). Anti-transferrin receptor scFv-
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immunoliposomes is another novel variation in clinical
practice against tumor (Liang et al., 2002).
Immunoliposomal based gene delivery has much impact
on managing hepatic carcinoma especially in transferring
conjugates (Ruiz et al., 2002). In the current scenario
immunoliposomes are one of the best delivery vehicles
for drugs and therapeutic genes in cancer management
(Kathy et al., 2007; John et al., 2002).
NANOPORES
Nanopores possess fine tiny hole and are
designed with pore forming protein or synthetic materials
like Silicon and Graphene. The prerequisite for the
design of nanopore is to create a channel for delivering
drugs, nucleic acid and for diagnostic applications (Ankit
et al., 2005; Dwaine et al., 2003). Available nanopores
are broadly categorized into biological nanopores made
up of protein and synthetic nanopores designed with
material like silicon, also referred as solid state
nanopores (Figure 3) (Andrei et al., 2009). Recently,
nanopore based DNA sequencing has become an
advanced tool in molecular biology due to the accuracy
and reproducibility of nanopores (Ian et al  
        

      
et al., 2010). In the case of design of solid state
nanopores, among numerous available materials, Silicon
as Silicon Nitride and Graphene are more common in
clinical practice (Chaitanya et al., 2011; Ping Xie et al.,
2012). There are very limited evidences available for
nanopore as a gene delivery tool for therapeutic
applications. However, nanopores are more commonly
employed as a novel tool for DNA sequencing in recent
times with high precision and accuracy (David et al.,
2009).
The nanopore based sequencing work is built on
the principle of enzymatic cleavage of DNA by a nucleus
enclosed within the nano-assembly. Since nucleic acid
possesses the charge on each of the nucleotide counted
immediately after cleavage and transferring into nano-
cleft (David et al., 2002), nanopore based sequencing
analyzes each single nucleotide during the process
(Liming et al., 2002). Additionally, nanopores do exhibit
the potential to carry a DNA cargo among various tissues
but is not clinically approved yet (Marijeta et al., 2003;
Masaya et al., 2002). Nanopores could be utilized as
carriers of cells or tissues rather than just nucleic acid, as
often required in diseases like Diabetes mellitus.
Conventional approaches for transplantation are often
limited by immune-complications. Beta cells  
transplantation, enclosed in nanopore, can be easily
carried out into the recipient with a minimum risk of
immuno-complications.
FULLERENES
Fullerene is a three dimensional structure of
carbon element in highly organized symmetry with 20
hexagonal rings and 12 pentagonal rings as shown in
figure 4 (Rania et al., 2007). Highly symmetric
configuration of fullerene is due to the bonding pattern of
each carbon atom bonded with other three Carbon atoms
by sp2 hybridization. (Koruga et al., 1996; Katsuhiko
Arigaa et al., 2007) Fullerene, a Carbon allotrope often
called as bucky ball was discovered in 1985 and the most
common fullerenes possess 7A
o
diameter and comprise
of 60 carbon atoms in icosahedrons (Steven et al., 2008).
Though, commonly known fullerenes are nanoscale
structures of the carbon atom referred to as homo-
fullerenes, another class of hetero-fullerenes with
nitrogen and boron are also reported recently. Fullerenes,
commonly found in nature, are reported to be present in
outer space and are believed to have provided seeds for
life on earth. There are several forms of these like bucky
ball cluster, nanotubes, megatubes, polymers,
nanoonions, linked ball and chain dimers, fullerene rings
etc. Homo-fullerenes, based on their three dimensional
shape and atomic structure, are further categorized as
C20, C26 (Dodecahedral), C60 and C70 fullerenes
(icosahedral) (Stephan et al., 2003; Atsushi et al., 2007).
Diverse structure and large space made
fullerenes to emerge as ideal vehicles for gene and drug
delivery. Studies performed in the last few years
confirmed the potential of fullerenes as novel vehicles in
gene delivery. In a study performed at University of
Tokyo, a successful delivery of insulin gene tagged with
EGFP was achieved for management of Diabetes mellitus
(Iroyuki et al., 2006). It was cationic termini
C
60
fullerene employed in-vivo for delivering EGFP
conjugated Insulin gene. The advantage of using of
cationic tetra amino conjugates was it offers high stability
of DNA against nucleases in tissues. Further, fullerene
based DNA delivery is becoming popular in DNA
vaccination. Additionally, in another study, binding
affinity of DNA with fullerene was evaluated with
fullerene tetra (piperazino) fullerene epoxide (TPFE) and
found enhanced under TPEE in comparison to fullerene
alone (Rui et al., 2010, Balaji et al., 2008). Fullerene
based delivery is aiming new high not only in the
delivery of nucleic acid (DNA) but also potent drugs
(Wang et al., 2009; Joseph Wang, 2005). From several
experiments conducted in vivo in animal models like rats,
it was inferred that there are no toxicity problems with
C60 and C70 but cannot be taken for granted with
fullerene derivatives, particularly with covalently bonded
chemical groups.
NANOTUBES
Nanotubes are refined fullerenes in the shape of
tubes comprising carbon atoms in systematic orientation
(Pauluhn, 2010). Carbon nanotubes were first discovered
in 1991 as tubular structures which can be single layer
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(Single walled Carbon Nanotube-SWCNT) or
multilayered (Multi walled Carbon Nanotubes-MWCNT)
(Figure 5) (Alberto et al., 2005; Davide et al., 2004).
SWCNT are 1-2 nm diameter while MWCNT are 2-
25nm diameter with 0.36nm space between two layers in
MWCNT (Zhuang et al., 2008). Contrary to other
Nanoconstructs, carbon nanotubes possess extra-ordinary
stability and strength that make them suitable to emerge
as ideal vehicles for drug and gene delivery (Marianna
and Mukasa, 2008). In order to achieve targeted delivery
and real time monitoring, carbon nanotubes can easily be
tagged with radio antibodies and nuclei (Alberto et al.,
2005). Another advantage with carbon nanotubes is that
they can be designed as water insoluble and water
dispersible as well, based on the requirement.
Additionally, half-life dependent renal and hepatic
clearance can be optimized by conjugation with
monoclonal antibodies and enhance targeted delivery of
candidate gene(s) and drug molecules (Xiangyang Shi et
al., 2009; Ritu Dhankhar et al., 2010).
When compared to other nano-designed
constructs for gene and drug delivery, carbon nanotubes
emerged as better vehicles with tremendous potential.
The most explored area in medicine using carbon
nanotubes is tumor management where drugs and nucleic
acids are selectively delivered (Yan-Yan Song et al.,
2009). In the year 2009, a successful targeting was done
in breast cancer cells using dendrimer modified carbon
nanotubes with excellent clinical results (Wuxu Z et al.,
2011). The enhanced application of the CNTs in delivery
of genes and drugs is due to their low toxicity and
immunogenicity (Thierry and Michael, 2002). Another
benefit using CNT as a delivery vehicle is it does not
interact with candidate biomolecules; hence offers free
and enhanced delivery of proteins, nucleic acids,
hydrophilic and hydrophobic drugs with the property
intact (Meital R and Ehud G, 2003). CNT based gene
deliveries are in clinical practice for many diseases such
as cystic fibrosis, muscular dystrophy, cancer and
diabetes (Thierry and Michael, 2002; Seyed et al., 2011).
Owing to these potentials, Multi Walled CNTs are
becoming more popular to deliver various biomolecules
such as protein, nucleic acid and drugs (Raffa et al.,
2011; Song et al., 2011).
QUANTUM DOTS
Quantum dots, a class of nanostructures of sizes
2-5nm can be designed in single or multiple layers of
material (figure 6) (Akiyoshi, 2004). The typical
structure of a quantum dot consists of the inorganic core
coated by biomolecules, either drugs or nucleic acids as
per requirements (Xiaohu et al., 2004). In recent times
quantum dots have been extensively used in medicine for
diagnostic and therapeutic purpose as well. Quantum dots
offer a large surface area for coating of therapeutic
molecules and deep penetration as are of nanoscale size
of 2-5nm (Robert et al., 2004; Igor et al., 2005).
Quantum dots further can be refined for targeted delivery
by conjugation with antibodies and radio nuclei (Aihua et
al., 2004). There are numerous methods available for
synthesis of quantum dots: colloidal synthesis, bulk
manufacturing and viral assembly are major ones
(Warren et al., 1998).
Among these fabrications, viral assemblies are
getting much importance in current scenario due to
control and ease in design. Another class of quantum dots
synthesized is cadmium free Quantum dots that show
preferred applications in household and food items
(Maksym et al., 2008). The potential of quantum dots in
gene delivery is getting magnified with time and
technology. With the ability to deliver therapeutic DNA,
quantum dots are now much engaged in the delivery of
si-RNA for gene silencing, a potential measure against
microbial infection especially viral, and molecular
imaging (Zhao et al., 2010). In the year 2011, a study
was performed to silence HPV16C6 gene in HeLa cells
by using l-arginine-functional-modified CdSe/ZnSe
Quantum dots (Hrvoje and Danuta, 2012; James et al.,
2003).
Studies have confirmed the role of quantum dots
in delivering si-RNA and the same was reconfirmed by
RT-PCR. This study made a breakthrough to fight against
pathogenic infections by a novel approach. Gene
silencing offers an excellent platform to block candidate
gene and is naturally found to occur among a few
organisms. Though gene Silencing was known for the
last few decades, its implications are still to be
understood carefully (Jin-Ming Lia et al., 2011). Now,
quantum dots based si-RNA delivery and inhibition of a
particular gene, may offer safe treatment of many
infectious diseases (Karl Deisseroth et al., 2006). With
this potential, quantum dots are ideal vehicles while
delivering therapeutic molecules in neuronal tissues
(James et al., 2009) as they often get damaged during the
delivery of therapeutics leading to more trauma (John et
al., 2004). External thrombolytics are often delivered in
conjugation with different quantum dots in the
management of cerebrovascular disorders (Antonio
Quintana et al, 2002).
DENDRIMERS
Dendrimers belong to a class of fine nano-scale
structures of (bio) molecules essentially branched with
highly definite geometry (Omar et al., 2003). A brief
description is available in figure 7 depicting the
molecular arrangement in a dendrimer. Unlike other
nanostructures, size of dendrimer depends on the extent
of branching and the molecule used for its synthesis (Kim
et al., 2002). A dendrimer consists of three sections:
inner core that allows conjugation to (bio) molecules,
outer surface with reactive terminal groups and in
between pincer facilitates binding of conjugated
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molecules on core (Wojciech Lesniak et al., 2005; Brian
and Norman, 2001). Often there is a little resemblance
between quantum dots and dendrimers as both look
similar superficially, but dendrimers are more symmetric
even with extensive branching than the quantum dots
(Matos et al., 2000). Additionally, dendrimers can be
designed in various classes such as charged, and based on
the total Charge of the molecule, dendrimers can be
anionic or cationic and offer efficient delivery to targeted
tissue or cell (Cameron et al., 2005). Branched periphery
of dendrimer offers ease in conjugation with radio nuclei
and antibodies and enhancing the efficiency of specific
delivery with real time monitoring of therapy (Gong Wu
et al., 2004). Dendrimers are synthetic nanoscale
constructs synthesized by divergent dendrimer,
convergent dendrimer, double exponential or mixed
syntheses (Jesse et al., 2008). Further, dendrimers are
categorized into many classes based on the type of
chemical molecules used in the synthesis (Loo et al.,
2005).
Dendrimers exhibit great potential of gene and
drug delivery in diverse tissues along with their
diagnostic applications especially in molecular imaging
(Stephen et al., 2001). Dendrimers have a long clinical
history as vehicles for drugs, recently transformed into
gene delivery particulates in cancer management (Parka
et al., 2000). The advantage of dendrimers as vehicles is
their easy uptake by cell by endocytosis and is quite safe
in the context of immune complications (Parka et al.,
2001). The potential of dendrimer as a carrier for
delivering DNA cargoes was carried out in mesenchymal
cells, a part of tissue regeneration and tissue engineering
     et al., 2010). The clinical
potential of dendrimer was confirmed by in-vivo study
carried out in conjugation with luciferase and GFP as a
reporter gene (Chia-Chun et al., 2006). In this study,
dendrimers tagged with gold nanoparticles were
transformed and efficiency evaluated with the expression
of reporter gene (Fumihiro et al., 2003). Dendrimers are
in routine use for gene delivery for neurological disorders
and deep ear disease (Perez-Martinez et al., 2012).
Another advantage of dendrimer as a vehicle is its
architecture that facilitates delivery of various
biomolecules for therapeutic and diagnostic purposes (
Loo et al., 2004).
NANOSHELLS
Nanoshells are an advanced combination of
biophotonics and nanotechnology, a hybrid technology
(Prodan et al., 2003). Nanoshells are spherical
nanoconstructs made up of a dielectric core which is
further coated with a fine metallic layer more often gold,
a cartoon in figure 8 (Hirsch et al., 2003). Silica core
nanoshells are 50 nm diameter while silver based are 10-
20 nm. The nanoshell based drug delivery has shown
tremendous benefits especially in cancer treatment and
credit goes to Professor Naomi J Halas for his inventions
in 2003 at Rice University (Jackson and Halas, 2001).
Nanoshells are purely synthetic constructs; the ones
coated with gold are ideal devices to carry potent drugs
and the same has been proven in cancer treatment by
using Nanoshells (Prashant et al., 2008; Dakrong et al.,
2008). Nanoshells loaded with potent therapeutic drugs
release on exposure to Near Infra-Red (NIR) radiation.
More often Nanoshells based drug delivery is employed
in cancer management where they exhibit cytotoxic
activity toward tumor cells after release of drug (Loo C,
et al., 2005).
Here, the mode of drug delivery is site specific
and targeted. Nanoshells utilize radiation energy to
diffuse drug across the biological membrane by
converting into thermal energy and raising local
temperature (Patrick et al., 2004). Despite all other
nanoscale designs, nanoshells release therapeutic cargoes
in desired tissue by exposure to Near Infra-Red (NIR)
radiations (Marites et al., 2008). This unique feature
provides ease of targeting of tissue and specific delivery
of therapeutic molecules (Xiaojun et al., 2007).
Nanoshells are ideal carriers for neuronal tissues and
tumors with minimum damage to healthy tissues (Rizia
Bardhan et al., 2011). Breast cancer characterized by
over-expression of HER2 gene can be turned off by
delivering si-RNA (Lee et al., 2005). Further, nanoshells
are ideal tools for molecular imaging using immunoassay
for particular tissues (Surbhi Lal; et al., 2008). Recent
studies have shown their potential mechanism of finding
metastasis and treatment subsequently (Vitaly et al.,
2011). Nanoshells also offer real time monitoring of
therapy and imaging by conjugation with
immunoglobulins, reporter genes and radio nuclei (Levy
et al., 2010).
NANOBUBBLES
Like nanoshells, nanobubbles are spherical nano
structures efficiently employed in managing cancer (Phil,
2003). Nanobubbles exist as fine mist/emulsion droplets
which are soluble at room temperature and are converted
into micro-bubbles upon heating (Ghaleb et al., 2008).
These are hollow droplets in contrast to nanoshells that
have a solid core (Figure 9). Another difference between
nanoshells and nanobubbles is that NIR radiation is
needed for release of encapsulated drug from nanoshells
while in the case of nanobubbles, ultrasonic waves drive
the release of drug (Natalya et al., 2007). In recent times,
intra capillary clot dissolution by sonothrombolysis
technique using nanobubbles is becoming more popular.
In addition, Nanobubbles using ultrasonic waves
have a high potential as diagnostic tool especially in
detecting cancer and tumor metastasis (Ekaterina et al.,
2012b). Nanobubbles are unique nanoscale architectures
specially designed for cancer management at the
molecular level (Ekaterina et al., 2012a). Moreover,
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Nanobubbles are currently employed in clot removal and
dissolution in vascular pipeline of deeper tissues using
ultrasonic wave called as sonothrombolysis. Very recent
studies carried out at Rice University demonstrated
plasmonic Nanobubbles based killing of diseased cells
including tumor cells (Carrie, 2010). In this study, heated
gold nanoparticles easily identified tumor cells and
released potent toxic drug leading to cell death (Usuke et
al., 2009). In another study, nanobubble mediated IL2
gene delivery and subsequent expression leading to cell
death was achieved with enhanced selectivity
(Lukianova-Hleb et al., 2010). Here, IL2 gene expression
led to immunological activation for targeting cancer cells
(Ranjita et al., 2010). With the known potential of
nanobubbles in cancer therapy, tissue regeneration
attempt has been made in the last few years using
nanobubbles as carriers for regulatory molecules (Brian
and Natalya, 2011; Castor and Trevor, 2005).
NANOSOMES
Nanosomes are phospholipid based double
membrane nanostructures and possess a close
resemblance with liposome, a nanosome cartoon in figure
10. The basic difference between a liposome and a
nanosome is the arrangement of phospholipid molecules
and more often the pattern of drug loading (Jasmina et
al., 2007). Another difference between the two is the way
they are synthesized: the supercritical fluid technology.
In a nanosome, the fluids first dissolve raw phospholipids
and then compress them. (Anup et al., 2010). More often
nanosomes are single-bilayer phospholipids while
liposomes are double/multiple layered. Nanosomes often
conjugated with photo-catalysts generate free radicals as
reactive oxygen and destroy targeted tissue under
exposure of light (Yasutaka et al., 2011). The
physiochemical properties of nanosomes such as size,
surface area, shelf life, half-life, all depend on the way it
gets manufactured and supercritical fluid used (Jose et
al., 2011). Nanosomes offer delivery of a variety of
biomedicals like hydrophilic recombinant proteins and
nucleic acids with hydrophobic drugs like anticancer
drugs and anti HIV drugs (Zarena and Sankar, 2011;
Philip, 2008).
Nanosomes are in routine use for delivering
nucleic acid for therapeutic purposes and inhibition of
expression of genes leading to pathogenicity (Chandra et
al., 2012). In the year 2012, a study has confirmed the
potential of magnesium as carrier for si-RNA for
inhibition of hepatitis viral gene expression. The
significance of the study can be evaluated as nanosomes
used in the study were able to deliver multiple si-RNA
leading to blockage of replication instead of a single
gene. This is going to be a breakthrough in fighting
against a different variant of hepatitis causing life
threatening consequences (Kathryn et al., 2012;
Surendiran et al., 2009). With the amazing potential of
nanosomes as delivery vehicles, many novel variants
have been designed to improve the efficiency (Frank et
al., 2010). Nanosomes in conjugation with high pressure
homogenization offer delivery of therapeutic molecules
for deeper tissues and neuronal tissues in particular (Sion
et al., 2009). Further, lipid based magnesium is in routine
use of nucleic acid delivery to skin disorders. In the study
(Pankhurst et al., 2003), UV light exposed skin cells were
targeted to reduce negative impact of high energy
radiation by delivering si-RNA.
MAGNETIC NANOPARTICLES
Magnetic nanoparticles are nanoparticles
conjugated with materials exhibiting magnetic properties
(An-Hui et al., 2007). Commonly used materials to
design magnetic nanoparticles are iron, nickel, cobalt and
other chemicals with the innate ability of magnetism (Lee
et al., 2006; Akira et al., 2005). Based on design,
magnetic particles can range 2-500 nm with immense
potential in therapeutics and diagnostics. However,
among several magnetic nanoparticles available,
paramagnetic iron oxide nanoparticles are often used for
imaging called as magnetic resonance imaging
(Taeghwan, 2003). Recently, in-vitro diagnosis trials of
breast cancer were done with paramagnetic iron oxide
nanoparticles conjugated with HER2 antibodies which
are unique for breast cancer cells (Yang et al., 2010).
Magnetic nanoparticles often face stability problems
especially internalization by macrophages and this could
be solved by treatment with lovastatin. Plasma opsonins
based stability problem can be resolved by treatment with
decoys (Duguet et al., 2006)
In recent times, magnetic nanoparticles
especially microcrystalline iron oxides are employed to
understand the molecular etiology in respect of brain
tissues with minimum risk and high diffusion across
biological barriers. Magnetic nanoparticles (MNPs) have
shown their potential for delivering DNA and other
biomolecules for diagnostics, delivering enzymes and for
therapies in recent times (Berry and Curtis, 2003).
Diagnostic application of MNPs became an essential part
of current medicine and simultaneous delivery of
therapeutic genes for anticancer therapy and
transplantations (Duguet et al., 2006). MNPs are ideal
tools for the imaging of the brain and neuronal tissues by
using microcrystalline iron oxide (MIONs) (Zhang et al.,
2008). MINOs based imaging of brain and other parts of
neuronal tissues avoids the chance of damage often
caused by conventional surgical methods (Beata
Chertoka et al., 2008). Besides these applications, MNPs
are ideal nano tools as they offer controlled and sustained
drug delivery essential for many disorders (Kim et al.,
2006). Further, MNPs tuned by conjugation with other
nanostructures such as Fullerene-tagged MNPs and
charged MNPs showed improved efficiency in the
delivery of biomolecules (Giri et al., 2005). Another
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study in cancer therapy especially breast cancer, where
Luteinizing hormone conjugated with antitumor drug was
targeted to LHRH receptor for delivery of drug was
completed with enormous success (Carola et al., 2006).
NANO-CONSTRUCTS IN CLINICAL PRACTICE
First ever implementation of successful gene
delivery as part of therapeutic medicine was the
management of Severe Combined Immunodeficiency
Syndrome (SCID) (Herzog, 2010). In the year 1990, viral
vehicle delivery of Adenosine Deaminase (ADA) led to a
revolution in the field of medicine. SCID treatment was
the first successful trial for delivering therapeutic genes
and Gene Therapy too (Salima et al., 2003). Yet, as it
was a virus based vehicle to deliver DNA cargo, many
complications were encountered in subsequent trials in
other diseases (Michael et al., 2004). This led researchers
and scientists to find novel tools for safe and efficient
delivery of therapeutic drugs and nucleic acids.
Numerous viral vehicles were designed genetically to
achieve the goal and have shown promising result but
with enormous complications (Schaffert and Wagner,
2008). Era of non-viral vehicles came into the picture
with the emergence of material science at fine level i.e. at
Nano Scale (Aliasger et al., 2003). In very short span of
time of 20 years, a number of non-viral delivery systems
established their potential as carrier molecules for
delivering therapeutic molecules. With the exponential
increase in nanoscale technology, the design and
development of various fine architectures such as
nanoparticles, nanotubes, nanofibers, fullerenes, quantum
dots, nanobubbles etc. became very much possible (Tong
and Nemunaitis, 2008; Volker Wagner et al., 2006).
The list of nano-scale devices approved by FDA
in routine use of delivering drugs is longer than the FDA
approved nano-architectures for gene delivery
(Farokhzada and Langerb, 2006). Nano-architectures for
management of tumors like lung and ovarian cancer as
albumin bound taxane particles and liposomal
doxorubicin respectively are in clinical use since one
decade (Kessinger et al., 2009). Albumin bound taxane
particles manufactured by Abraxis Oncology Laboratory
has the brand name of Abraxane while Doxil from Ortho
Biotech is branded marketed product for liposomal
conjugate of Doxorubicin (Mark et al., 2008).
Additionally, liposome based Amphotericin B nano-
architecture is available for systemic fungal infection as
AmBisome from Astellas Pharma (Kawasaki et al.,
2005). Besides these approved products, numerous nano-
constructs are in different phases of clinical trial studies
(some enlisted in table 1) and hopefully, very soon they
would be available for commercial use. Iron oxide
nanoparticles for molecular imaging from Dendritic
Nanotechnologies Inc. is in Phase III and has shown
promising results in preclinical study in vital tissues like
heart and brain (Sanjib Bhattacharyya et al., 2012).
Numerous other nano-designs for cancer
management are in different stages of clinical trials like
dendrimer based microbicide gel for HIV protection
(Phase I). In late 1990s nanoscale architectures entered
into medicine first as tools for molecular diagnosis and
later as carriers of therapeutic molecules like proteins,
nucleic acids and drugs (Kim et al., 2010). With their
robust applications, existing molecules were refined to
enhance efficiency and thus several novel designs were
invented (Annalisa Palmieri et al., 2007). In the last
decade, many of nano scale architectures got approved by
the FDA and the number is increasing rapidly (Yechezkel
and Doxil, 2012). FDA approved nano scale architectures
are in routine use in medicine as tools for molecular
diagnosis, delivering of therapeutic protein drugs and
nucleic acids (Vinod Labhasetwar, 2005). A complete
overview of nano scale architectures in medicine is
summarized below with their potential and future
prospects in medicine.
SIGNIFICANCE OVER EXISTING VEHICLES
Non-viral and especially nanoconstruct based
gene delivery provides numerous benefits over virus
based options. The most important finding of
nanoconstructs over viral vehicles is their safety (Haibin
et al., 2002). Viral based vehicles are often associated
with diseases and secondary infections with course of
time or post therapy periods (Schatzlein, 2001). Another
advantage of nanoconstruct based vehicles is their higher
shelf and half-life and less immune-complications as
compared to viral based vehicles (Glasgow et al., 2006).
Also, nanoconstructs can carry larger size of the
candidate nucleic acid often needed for many diseases
(Thomas and Klibanov, 2003). Additionally,
nanoconstructs can be designed in a variety of vehicles
based on requirements as discussed in this study which is
quite difficult with other tools for gene delivery
(Lungwitz et al., 2005). In the case of nanoconstructs,
their manufacturing, application and storage conditions
are safe and easy to furnish compared to virus based
vehicles (Lu et al., 2003). Further, conventional tools do
not provide real time monitoring of gene delivery and
expression (Colin and Leonard, 1998). New generation
nanoconstructs provide dual benefits: one targeted
delivery and the other, real time monitoring as often
conjugated with a marker (fluorescence or radio nuclei)
(Gerrit Borchard, 2001).
Real time monitoring defines the extent and fate
of therapy and molecular trafficking (Niidome and
Huang, 2002). The unique design of nanoconstructs and
their ultra-fine size allow for deeper penetration beyond
biological barriers and subcellular trafficking (Maureen
et al., 2001). Viral vehicles and conventional physical
methods for gene delivery often face stability problems
against immunological and hydrolytic enzymes in tissues
and blood plasma and this culminates in the failure of
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delivery and therapy (Mannello and Gazzanelli, 2001).
Nanoconstructs have shown tremendous stability against
hydrolytic enzymes and immune system with high rate of
gene delivery. In the case of management of neurological
disorders like Multiple Sclerosis that needs delivery of
gene into central nervous system beyond the blood brain
barrier, conventional delivery tools, due to poor
infusibility and risk factors associated with viral vehicles,
are often a failure (Mohamed, and Dennis, 2008) while
nanoconstructs efficiently cross the blood brain barrier
and hence deliver candidate genes with minimum
obstacles (Christine and Barrett, 2003). Nanoconstructs
do offer dual applications: diagnosis (Molecular imaging)
and therapeutics (Delivery of drugs and biomolecules-
protein and nucleic acid) (Haerim et al., 2007). Most of
the nanoconstructs with their immense potential as
delivery vehicles are also employed as diagnostic tools at
molecular level, a much difficult task in the case of viral
vehicles (David et al., 2007). Diversity in nanoconstructs
and the ease of their fine refinement as per requirements
make them stand as a first choice for delivery of both, the
drug and the biomolecules. Finally, Gene therapy often
struggles with ethical issues and are more distinct while
using viruses as delivery vehicles which need genetic
modulations against ethical protocols. Nanoconstructs are
often free from ethical violations and are in clinical
application for the delivery of numerous potent drugs and
biomolecules. For these reasons, nanoconstructs are the
preferred choice for delivery of drugs and biomolecules
with a higher success rate and least complications
(Gaetano R et al., 2000).
Table 1. Nanotechnology based products in clinical trials.
Product
Name
Description
Current status
Application
CALAA-01
Cyclodextrincontaining si-
RNA
Delivery nanoparticles (~50
nm) based on Calando's
RONDEL technology
Phase I
For the management of
various cancers
INGN-401
liposome FUS-1
Phase I
For the management of
metastatic, non-small
cell lung cancer
Aurimmune
(CYT-6091)
Colloidal gold nanoparticles
coupled to TNF and PEG-
Thiol (~27 nm)
Phase II
For the management of
Solid Tumors
SGT-53
p-53 liposomes
Phase I
For management of
various Solid Tumors
NB-00X
Nano- emulsion droplets (~200
nm) based on Nano-Stat
Technology
Phase II
Herpes labialis caused
by herpes simplex I
virus
Auro-Shell
Gold-coated silica
nanoparticles (~150 nm)
Phase I
For management of
various Solid tumors
Fig 1. A diagrammatic illustration of liposome; A- an
amphipathic phospholipid molecule, B- arrangement of
phospholipid molecules and C- special arrangement of
phospholipids as liposome.
Fig 2. A diagrammatic illustration of immune-liposome:
liposome conjugated with antibodies.
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Fig 3. A diagrammatic illustration of nanopore spanning
biological membrane, B Graphene nanopore for DNA
sequencing.
Fig 4. Diagrammatic illustration of carbon fullerene:
Homo-Fullerene,C70.
Fig 5. Diagrammatic illustration of carbon nanotube A- Single
Walled Carbon Nanotube (SWCNT), B- Multi Walled Carbon
Nanotube (MWCNT)
Fig 6. Diagrammatic illustration of quantum dot, A-
typical structure of quantum dot core conjugated with
chemical, B- Quantum dot conjugates with biomolecules
protein and DNA.
Fig 7. Diagrammatic illustration of Dendrimers, A- Structural
arrangement, B- overview of mature Dendrimer.
Fig 8. A typical cartoon of nanoshell, core coated with
gold sheet with tiny dipole offer ideal platform for
delivery of drugs and biomolecules.
Fig 9. Diagrammatic illustration of Nanobubbles emerging out
from the tumor cell membrane after exposure with ultra-sonic
waves.
Fig 10. A diagrammatic illustration of nanosome consists
of single bilayer of phospholipid molecules, ideal for
hydrophilic biomolecules like protein and nucleic acid.
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CONCLUSION
The Nanoconstruct based gene delivery holds
great potential in current medicine for managing
therapeutics at the molecular level with enhanced
efficiency. Wide range of nanoconstructs and their wide
spectrum of applications led to a new era of medication
which is not only safe but also offers a permanent cure of
many life threatening diseases and disorders. Rapid
improvement of technology in material sciences allowing
the revolution in biological sciences promotes better
understanding of the molecular etiology of cells, their
abnormalities and treatment subsequently. Another area
of health care system i.e. molecular diagnostics is
exponentially      
nanoconstruct based diagnostic tools. The number of
clinically approved nanoconstructs is growing
     
that made possible successful cure of numerous life
threatening diseases considered as major trauma in the
last century. With the potential applications of nano-
scale designs as therapeutics and DNA carriers, in the
recent past, gene silencing by nanoconstruct delivered si-
RNA became more popular. There is a tremendous scope
and requirement for such tools to overcome viral, fungal
and bacterial diseases in human, animals and plants.
Nanoshells and nanosomes are in clinical application for
delivering si-RNA against pathogens and inborn errors.
Breast cancer often characterized by over expression of
HER2 gene turns off by si-RNA delivered by Nanoshells.
Studies also confirmed that hepatitis viral replication can
be blocked by si-RNA and thus provide safe and durable
treatment. More emphasis is given in current material
science to design and improve efficiency of nanoscale
designs for use in medicine (therapeutic and diagnostic)
and agriculture.
FUTURE PROSPECTS
Though potential of Nanoconstruct based
medicine and gene delivery is widely accepted,
implementation is hindered for a few reasons. Major
challenges for nano scale technologies in biological
Sciences are their approach and cost factor. Still this
advanced technology is not approachable as compared to
traditional medicine. Hence more attention is required to
make this amazing technology available to the individual
subject. Another area which has to get explored in
reference to nano-product as a tool for gene therapy is
public awareness and education. Ethical obstacles are
much more frequent in gene therapy using either non-
viral or viral methods. These are to be revised again in
order to potentiate technology for the benefit of human
health and civilization.
ACKNOWLEDGEMENTS
We would like to convey our sincere thanks to
the Management and the Principal, R.V.R. & J.C.
College of Engineering, Guntur, Andhra Pradesh, India
for their encouragement.
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... The comparison between the immune system of Vertebrates versus in-vertebrates should be on the structural level but must be on its functionality. For example, our immune system is a complete collapse in the case of novel SARS-CoV-2 infections that raise a question immediately do we are really superior over lower animal invertebrates encounter daily such infections [1][2][3][4][5][6]. ...
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Humans evolved with a unique immune system consist of innate and adaptive immunity. Both innate and adaptive immunity comprised of different types of cells, immune players, and associated with different functions. In higher animals, including humans, innate immunity derived from birth hence called inbuilt immunity. On the contrary, adaptive immunity called acquired immunity is gain due to the environment. Vertebrates are ranked as superior over invertebrates on the classification table due to many reasons, including the presence of vertebra and spinal cord. Invertebrates often considered as lower animals due to poorly developed organs, lack of vertebra and spinal cord. Similarly, the immune system and set of immune organs in a lower organism are entirely different and primitive on the basis of the organization. Like humans and other vertebrates, the invertebrates do not have a distinct clas-sification/setup of innate and adaptive immunity. In human and higher vertebrates, innate and adaptive immune systems are consisting of different cells; immune players such as cytokines, anti-bodies, and chemokines, etc. enable crosstalk between innate and adaptive immune systems. On the contrary, invertebrates' immune system is consists of unique cells and potent biomolecules; enzyme , peptides, and proteins. These molecules provide a complete defense to the animal via several mechanisms. The question we raise here does the classification superiority of humans, and other vertebrates over in vertebrates are justified in terms of immunity. How a complex and evolved immunity system mostly in human often fail to fight against infections while lower animals with the primitive immune set can survive even in hearse conditions. Apart from organization setup in humans and higher verte-brate's immunity can be further classified based on function such as cell-mediated and Humoral immunity. Cell-mediated immunity provides protection using different cells such as white blood cells (WBC), Macrophage, Phagocytes, Dendritic cells and T cell (CD4 and CD8), etc. Humoral immunity is a function of B cells, memory cells and antibodies. The immune response in human and other higher vertebrates consist of activity of different cells and immune players for identification of the foreign substance, presentation to immune system followed by killing/removal. The effective identification and clearance of foreign objects depend on synchronization between immune cells and players of both innate and adaptive immunity. On the contrary, lower animals, i.e. invertebrates, do have a more straightforward mechanism of identification and clearance of foreign objects that ultimately depends on endogenous short peptides, proteins, and enzymes. The more straightforward sets often provide ease in synchronization of immune players in lower animals reported more effective and robust. There is growing evidence that enzyme and protein as part of the immune system in the lower animal are highly dynamic and do possess promiscuity at the substrate and catalytic level. Enzyme promiscuity is capacity often describe for broad substrate affinity and catalytic diversity. The short peptides, proteins, and enzymes present in lower animals; invertebrates are highly promiscuous and capable of catalyzing multiple biochemical reactions. Earthworm, a classic invertebrate model for immune comparison , studied tremendously in the last couple of decades. The animal habitat is an environment rich in infections and invading elements. The presence of promiscuous and potent biomolecules including enzyme; serine protease provides a complete defense to the animal. The coelomic cavity of animals is rich in many short peptides, and other bioactive molecules offer extended protection to a wide range of infections. The key molecules are Fetidin 1, Fetidin 2, Lysenin, Eiseniapore, Coelomic Cytolytic Factor (CCF), and Lumbricin I. These molecules exist in isoforms and vary in size among different species of earthworm. The isoforms of these molecules provide extended support in defense mechanisms. There are growing research findings suggested protease present in coelome of earthworm are generally exist in isoforms. Each isoforms possesses a broad substrate affinity and catalytic diversity. This can be compared with VDJ recombination in B cell for the diversity of anti-bodies. There is another advantage in having promiscuous and iso-forms of series protease in coelome of animal, i.e. a large number Citation: Mahendra Kumar Verma and MV Raghavendra Rao. "Mapping Immunity; Vertebrates Versus Invertebrates". Acta Scientific Biotechnology 1.8 (2020): 42-43.
... The side effect and toxicity of CNTs depends on several factors including the method of synthesis, purification, and presence of the functional group. 132 There is an extensive research work underway to surmount these limitations of CNTs and hope in future CNTs will be playing a vital role in biomedical engineering. However, modulating CNTs physiochemical properties and translate them into drug cargoes remain a major challenge. ...
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The main objective in gene delivery system is the development of efficient, non-toxic gene carriers that can encapsulate and deliver foreign genetic materials into specific cell types To perform gene transfer two types of vectors are available, (i) viral vectors (ii) nonviral vectors. Among the vectors, nonviral vectors are proved less toxic and safe compare to viral vectors through clinical trial. No single vector is proved suitable for every gene transfection experiment. Cationic polymers such as polyethylenimine (PEI) and poly L-lysine (PLL) and their complexes are experimentally established as non-viral vector with higher transfection efficiency. Identifying the barriers for transfection and the possible solution, the stability improvement research is needed for better clinical performance of cationic polymers. In future it is also possible to find new conjugates as cationic polymers bearing many properties, suitable to bound with DNA and penetrate cell. Considering all these properties, these works reviews the most recent studies highlighting cationic polymers used in nonviral gene delivery systems.
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Ex-vivo gene therapy can entail either the replacement or the addition of genes. In gene addition therapy, a therapeutic gene is inserted directly into the host genome, with the abnormal gene remaining intact. In gene replacement therapy, the genome is modified directly. Homologous recombination technology can be used to perform many of these kinds of gene correction. In the past, gene correction therapy has been hampered by the low efficiency of the recombination event. However, recently engineered zinc finger nucleases (ZFNs) were found to have the ability to successfully stimulate homologous recombination by inducing double-strand breaks at specific DNA sites. Another class of enzyme, the transcription activator-like effector nucleases (TALENs), provides an efficient alternative means to induce specific DNA double-strand at breaks. Meanwhile, newly developed gene correction methods using stem cells and induced pluripotent stem (iPS) cells have made gene therapy more feasible in clinical practice. Cells are taken from patients, harvested, and transformed through induction into stem cells, which have the potential to differentiate into a variety of mature cells types for transplant. Further research is needed to develop gene therapy, which may be used in tandem with embryonic and induced pluripotent stem cell therapy, especially to repair preexisting mutations that may be passed on in iPS cells.
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Dendrimers are repeatedly branched and roughly spherical large molecules. They can be used in various medical applications, such as anticancer polymeric nanomedicines and nanocarriers, gene carriers and vectors in gene delivery, contrast agents for molecular imaging and vaccines against infectious diseases and cancer. The highly branched, multivalent nature and molecular architecture of dendrimers make them ideal tools for a variety of tissue engineering applications. This book describes different categories of dendrimers, their biomedical and physico-chemical applications as well as convergent and divergent syntheses, click chemistry and ligation strategies. It is a rich source of information for researchers in biochemistry and pharmacology working on drug development as well as for organic chemists who are engaged in synthesis of dendrimers.
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Background. Stable transfection of Green Fluorescent Protein (GFP) has been widely used as a marker for both monitoring protein expression and direct observations of cellular dynamics. Despite the proven utility of GFP as a marker, the appropriate conditions of stable transfection, like selection of optimal concentration of G418 and the effect of passage number on fluorescence emission time has not been fully described for stable transfection of kanamycin/neomycin resistant pIRES-EGFP to Chinese Hamster Ovary (CHO) cells by activated dendrimers. Methods. To address the issues and the establishment of GFP expressing CHO cells for further experiments, we transfected the cells with pIRES2-EGFP by using activated dendrimers. Transfected cells were selected by eluding the reminder population by exposing to G418 (500 μg/ml for 3 weeks). Selected transfected cell strains were imaged by using confocal microscopy. Results. After 48 hours following transfection, 5% of cells were transiently transfected and the emitted green fluorescence faded within 4 days. Following G418 treatment for 3 weeks, 10% of the survived cells expressed GFP with a fluorescence intensity several folds larger than that in the control cells. GFP expressing cells had complex morphology which is different from the typical fusiform shape of CHO cells. A part of stably transfected cells showed some important changes in their morphology even though no green fluorescence was detected. Conclusions. This study demonstrates that fluorescent proteins can succesfully be expressed in mammalian cells for multiple purposes. Our future goal is to obtain stable GFP expression in different cell lines to be used for different experimental purposes.
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Over the past years, the gelation of aqueous solutions by low molecular weight (LMW) compounds has become an area of increasing interest, owing to developments in the field of LMW organogelators. Until recently, LMW hydrogelators were found only by serendipity, nowadays rational design as well as application of LMW hydrogelators has become feasible. As a consequence, an increasing number of responsive and functional LMW hydrogels are reported, offering great prospects for diverse applications including drug delivery and smart materials. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2005)
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Magnetic nanoparticles have been proposed for use as biomedical purposes to a large extent for several years. In recent years, nanotechnology has developed to a stage that makes it possible to produce, characterize and specifically tailor the functional properties of nanoparticles for clinical applications. This has led to various opportunities such as improving the quality of magnetic resonance imaging, hyperthermic treatment for malignant cells, site-specific drug delivery and the manipulation of cell membranes. To this end a variety of iron oxide particles have been synthesized. A common failure in targeted systems is due to the opsonization of the particles on entry into the bloodstream, rendering the particles recognizable by the body's major defence system, the reticulo-endothelial system. This review discusses each of the above bio-applications of such magnetic nanoparticles and details some of the main recent advances in biological research.
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Viscosimetric, hydrodynamic and conformational properties of dendrimers and dendrons were studied. The molecular conformation in solution and the melt that controls the efficacy in biocides, gene transfer and catalyst supportive applications was investigated with coarse grained simulations and extensive experimentation of x-ray light and neutron scattering and rheology. The radius of gyration was calculated from static scattering measurements while hydrodynamic radius was determined by light scattering and other diffusion measurements. Larger ratios were observed for polymers in solution.