**8. Advancements in nano-enabled therapeutics for HIV management**

Human immunodeficiency virus (HIV) is a deadly infectious disease worldwide [48–50]. The World Health Organization confirms 0.35 million HIV-infected people. Apart from them, 28 million people are eligible for antiretroviral therapy (ART and only 11.7 million could afford antiretroviral (ARV) drugs. But the ultimate challenge in highly active ART is the elimination of HIV-1 reservoirs from the peripheral nervous system and central nervous system (CNS) [51]. The integration of HIV-1 genome with host genome causes viral latency in the

**109**

*Using Microbubbles as Targeted Drug Delivery to Improve AIDS*

periphery and in brain. However, the inability of ART to penetrate the blood– brain barrier (BBB) after systemic administration makes brain as one of the most dominant HIV infection reservoirs [48]. Recently, dual therapy, i.e., an optimized cocktail of two ARV drugs, has been introduced to manage HIV infection by Kelly et al. [52]. The authors claimed that the dual therapy containing tenofovir (TEF) exerted more therapeutic advantages than triple therapy. Furthermore, the selection of appropriate drug according to the patient condition is very essential because this therapy may reduce virologic efficacy in HIV-infected patient while lowering CD4 counts per high pre-ART HIV-1 RNA level. This report stated that new nanoformulations (NFs) of LA cabotegravir (CTG) and rilpivirine (RPV) may have bright future aspect for HIV therapeutics. This viable dual therapy is useful to manage ART options and performance, which lowers the costs and the globally unmet needs of pill-fatigued and adherence-challenged individuals [52]. The pharmacologic profile of CTG has great potential for the treatment and prevention of HIV-1 infection. This drug has half-life of 40 days and at a low dose showed therapeutic action, so that monthly and bimonthly oral administration in the form of tablet would be enough to control HIV infection [53]. As the significant advancement made in antiretroviral drug for HIV very few efforts have been developed for effective anti HIV vaccine [54]. To cure neuro HIV the inability of effective anti HIV therapeutic agents to cross the complex integrity of the BBB is the major challenge, so neuro HIV is incurable in the brain. Specific receptor binding, focused ultrasound, microbubble assisted focused ultrasound and magnetic field based approaches have been demonstrated to open the BBB for the delivery of therapeutic agents. Due to bigger in size receptor-functionalized therapeutic cargos affect efficacy whereas an externally stimulated approach results in transient BBB opening, which may also allow the delivery of unwanted

Fluoresce activated cell sorting (FACS) is a standard method for diagnosis of AIDS but having high cost and beneficial only in area where large number of HIV patient resides. The second standard technique is Magnetic activated cell sorting (MACS) used for CD4 cell counting. It involves mixing of sample with magnetic beads which get attached to anti bodies. AIDS can be monitored by using the microbubbles which require no expensive equipment and low cost as compare to above methods. In this technique for separation of CD4 T cell lymphocyte from whole blood cell microbubbles are used. By mixing target specific antibody with microbubbles, the microbubbles get float on surface and provide eminent contact between microbubbles and target cell so that target cell attach to microbubbles

Ultrasound in presence of microbubbles increases plasmid transfusion efficiency *in-vitro*. Microbubbles form pores upto 100 nm by cavitation mechanism which is having short half life. Loading microbubbles with nucleic acid and /or disease targeting ligand may improve efficiency and specificity. Generation of reversible pore in the plasma membrane due to sonoporation increased plasma membrane permeability to marker compounds. Recent studies state that the effect of low frequency 20 Hz by ultrasound on uptake of fluorescent dye calcein having molecular weight 623 Da and radius 0.6 μm into mouse increase cavitation. This data explains that cavitation occur during insonication influence the membrane permeability. Practical and theoretical experiment on microbubble state that the rapid bubble expansion collapse and subsequent shock wave formation can generate shear forces which disturb cell membrane integrity and increased permeability. Further, the geometry of microbubble collapse is itself influenced by adjacent cell membrane like microjet of the surrounding fluid which in transfection medium

while non-targeted cell at the bottom side due to gravity [56].

contains exogenous nucleic acid may get injected in cell [57].

*DOI: http://dx.doi.org/10.5772/intechopen.87157*

agents to the brain [55].

#### *Using Microbubbles as Targeted Drug Delivery to Improve AIDS DOI: http://dx.doi.org/10.5772/intechopen.87157*

*Pharmaceutical Formulation Design - Recent Practices*

1.**Drug incorporation into microbubbles:** incorporation of drug molecule in the microbubbles in a following way (1) incorporation of drug molecule only within bubble, (2) incorporation of drug molecule within cell membrane, (3) attachment of drug molecule to microbubbles by covalent bonds, (4) attachment of drug molecule to microbubbles by ligand (ex avidin-biotin complex), and (5) incorporation of drug molecule in multiple layer of microbubbles.

In **Figure 3** by attaching a targeted ligand such as monoclonal antibody a targeted microbubbles are developed. These are specific for endothelial marker as microbubbles. To assess vascular pathology targeted ultrasound contrast

2.**Release of drug from microbubbles:** microbubbles on application of ultrasound undergo a process known as cavitation. Ultrasound causes the microbubble to burst or break. The body fluids begin to insonate on cavitation to create acoustic cavitation. After oscillating microbubbles produces increase small eddy, this increases the permeability of cell membrane and drug passes across the membrane. Microbubbles also release the drug by phagocytosis mechanism. **Figure 5** describe the delivery of drug through fusion mechanism in which the phospholipid microbubble fuse with phospholipid bilayer of cell membrane and releasing of drug or gene into the cytoplasm of cell membrane. By this mechanism the gene get directly transfer to the nucleus of the cell [44–47].

**8. Advancements in nano-enabled therapeutics for HIV management**

Human immunodeficiency virus (HIV) is a deadly infectious disease worldwide [48–50]. The World Health Organization confirms 0.35 million HIV-infected people. Apart from them, 28 million people are eligible for antiretroviral therapy (ART and only 11.7 million could afford antiretroviral (ARV) drugs. But the ultimate challenge in highly active ART is the elimination of HIV-1 reservoirs from the peripheral nervous system and central nervous system (CNS) [51]. The integration of HIV-1 genome with host genome causes viral latency in the

agent are used ex: P-selectin, ICAM-1, GpIIb/IIIa, the αv integrins.

Microbubbles are able to cross the BBB through above process.

**108**

**Figure 5.**

*Drug release from microbubbles by cavitation.*

periphery and in brain. However, the inability of ART to penetrate the blood– brain barrier (BBB) after systemic administration makes brain as one of the most dominant HIV infection reservoirs [48]. Recently, dual therapy, i.e., an optimized cocktail of two ARV drugs, has been introduced to manage HIV infection by Kelly et al. [52]. The authors claimed that the dual therapy containing tenofovir (TEF) exerted more therapeutic advantages than triple therapy. Furthermore, the selection of appropriate drug according to the patient condition is very essential because this therapy may reduce virologic efficacy in HIV-infected patient while lowering CD4 counts per high pre-ART HIV-1 RNA level. This report stated that new nanoformulations (NFs) of LA cabotegravir (CTG) and rilpivirine (RPV) may have bright future aspect for HIV therapeutics. This viable dual therapy is useful to manage ART options and performance, which lowers the costs and the globally unmet needs of pill-fatigued and adherence-challenged individuals [52]. The pharmacologic profile of CTG has great potential for the treatment and prevention of HIV-1 infection. This drug has half-life of 40 days and at a low dose showed therapeutic action, so that monthly and bimonthly oral administration in the form of tablet would be enough to control HIV infection [53]. As the significant advancement made in antiretroviral drug for HIV very few efforts have been developed for effective anti HIV vaccine [54]. To cure neuro HIV the inability of effective anti HIV therapeutic agents to cross the complex integrity of the BBB is the major challenge, so neuro HIV is incurable in the brain. Specific receptor binding, focused ultrasound, microbubble assisted focused ultrasound and magnetic field based approaches have been demonstrated to open the BBB for the delivery of therapeutic agents. Due to bigger in size receptor-functionalized therapeutic cargos affect efficacy whereas an externally stimulated approach results in transient BBB opening, which may also allow the delivery of unwanted agents to the brain [55].

Fluoresce activated cell sorting (FACS) is a standard method for diagnosis of AIDS but having high cost and beneficial only in area where large number of HIV patient resides. The second standard technique is Magnetic activated cell sorting (MACS) used for CD4 cell counting. It involves mixing of sample with magnetic beads which get attached to anti bodies. AIDS can be monitored by using the microbubbles which require no expensive equipment and low cost as compare to above methods. In this technique for separation of CD4 T cell lymphocyte from whole blood cell microbubbles are used. By mixing target specific antibody with microbubbles, the microbubbles get float on surface and provide eminent contact between microbubbles and target cell so that target cell attach to microbubbles while non-targeted cell at the bottom side due to gravity [56].

Ultrasound in presence of microbubbles increases plasmid transfusion efficiency *in-vitro*. Microbubbles form pores upto 100 nm by cavitation mechanism which is having short half life. Loading microbubbles with nucleic acid and /or disease targeting ligand may improve efficiency and specificity. Generation of reversible pore in the plasma membrane due to sonoporation increased plasma membrane permeability to marker compounds. Recent studies state that the effect of low frequency 20 Hz by ultrasound on uptake of fluorescent dye calcein having molecular weight 623 Da and radius 0.6 μm into mouse increase cavitation. This data explains that cavitation occur during insonication influence the membrane permeability. Practical and theoretical experiment on microbubble state that the rapid bubble expansion collapse and subsequent shock wave formation can generate shear forces which disturb cell membrane integrity and increased permeability. Further, the geometry of microbubble collapse is itself influenced by adjacent cell membrane like microjet of the surrounding fluid which in transfection medium contains exogenous nucleic acid may get injected in cell [57].
