Applications of Chitosan in Pulmonary Drug Delivery DOI: http://dx.doi.org/10.5772/intechopen.87932

relatively high, and the weak stability will lead to aggregations and interactions between the NPs. Thus, special surface modification will also be an effective way to

Kenneth A. Howard et al. had developed CS-based siRNA-loaded NPs for pulmonary RNA interference therapy [63]. The negatively charged siRNA was complexed by the positively charged CS to form the polyelectrolyte complex NPs. The particle size ranged from 40 to 600 nm. These NPs showed a rapid uptake (1 h) into NIH 3T3 cells. The NPs could mediate efficient knockdown of endogenous enhanced green fluorescent protein (EGFP) in both H1299 human lung carcinoma cells and murine peritoneal macrophages in vitro. Moreover, effective in vivo RNA interference was also realized in the bronchiole epithelial cells of transgenic EGFP mice after nasal administration. The results indicated that this kind of CS-based complex NPs has great potential in RNA interference therapy for systemic and mucosal disease.

A series of CS/fucoidan (CS/F) NPs had been designed and prepared as PDDS

(A) The preparation of the GM-loaded CS/F NPs. (B) TEM of the GM-C5F1 NPs. (C–E) Long-term stability

for gentamicin (GM) in Yi-Cheng Huang's study (Figure 2) [64]. The NPs presented a biphasic release feature, with a zero-order release of GM for the first 10 h, followed by a sustained release of up to 72 h, and the cumulative release value reached 99%. The GM-loaded CS/F NPs exhibited multiple antimicrobial capabilities against Klebsiella pneumoniae. The intratracheal administration of the GMloaded CS/F NPs displayed a higher area under the curve (AUC) and lower minimum inhibitory concentration ratio than the free GM that was administrated by intravenous injection. These results had showed that the CS/F NPs had superior antimicrobial efficacy and lower risk of systemic toxicity, and this GM pulmonary

delivery system has exhibited good potential for pneumonia treatment.

improve the stability of the NPs.

Role of Novel Drug Delivery Vehicles in Nanobiomedicine

Figure 2.

170

of the series of CS/F NPs.

Abdallah Makhlof et al. synthesized a thiomer derivative of glycol CS (GCS), which was coupled with thioglycolic acid (TGA) [65]. The NPs were prepared with GCS and GCS-TGA by ionic gelation for the pulmonary delivery of peptides. The NPs were positively charged and had sizes in the range of 230–330 nm. They also showed high calcitonin entrapment. After intratracheal administration, the mucoadhesion capacity of the GCS-TGA NPs was much better than that of nonthiolated NPs in rats. The toxic effect of the NPs with lung tissue was confirmed with negligible epithelial damage and toxicity. The NPs could enhance the pulmonary absorption of the delivered peptides, and the calcitonin-loaded GCS and GCS-TGA NPs showed efficient hypocalcemic effect. The GCS and its thiomer derivative could be used as potential PDDS for delivering peptides.

In another work, Adriana Trapani et al. had prepared CS and GCS NPs containing Lipoid S100 for the systemic delivery of low molecular weight heparin (LMWH) by pulmonary administration [66]. The NPs were prepared by ionotropic gelation method. The NPs were positively charged and with nanoscale size. Efficient drug entrapment and good mucoadhesive property could be achieved by using these NPs. The LMWH was effectively delivered into the lung by the aerosolization of the drug-loaded NPs. These results revealed the promising characteristics of the CS-based NPs, which were highly applicable for PDDS.

Tejal Rawal et al. designed a NP-based dry powder formulation of rifampicin (RFM) for achieving local and sustained targeting of anti-tubercular drugs to reduce dose and frequency [67]. The drug-loaded NPs were formulated by the ionic gelation probe sonication method. The optimized formulation had a small particle size of 124.1 nm with an entrapment efficiency of 72%. No toxicity was found by in vitro and in vivo experiments. The pharmacokinetic assay verified that the NP formulation would serve a better alternative because it could minimize the frequent dosing schedule than conventional dry powder inhalation and market formulation. The RFM-loaded NPs open new avenues for developing therapeutic interventions for lung tuberculosis.

## 3.2.3 CS-modified liposomes

Liposomes are primarily used as PDDS for the treatment of respiratory distress syndrome and other lung diseases [68]. The drugs carried by liposomes mainly have stable physicochemical properties and strong lipophilicity. Liposome-based drug carrier has been one of the research hotspots in pharmaceutics [69, 70]. At present, drug-loaded liposome preparations, such as for amphotericin B, daunorubicin, doxorubicin, cytarabine, and morphine, have been successively developed [71]. The liposome preparations of these drugs have many unique advantages compared with their common preparations. Liposome-based pulmonary administration has the advantages of rapid absorption, little irritation, good tolerance, high safety, controllable drug release, and improved bioavailability [72]. The encapsulation of the drug in liposomes affects the pharmacokinetic property of the drug and prolongs drug half-life. Moreover, 85% of the components of the alveolar surface are phospholipids, which can facilitate the liposomes accumulate and enter the lung tissue. Liposome-loaded macromolecules with low lipophilicity can significantly improve bioavailability, and liposomes can also reduce the side effects of the toxic drugs to normal tissues by pulmonary administration.

Marco Zaru et al. prepared CS-coated liposomes and used them as drug carriers for pulmonary delivery by nebulization [73]. Rifampicin (RFM) was loaded in the CS-coated liposomes with different lipid compositions. By coating with CS, the encapsulation efficiency of the liposomes increased slightly, and after nebulization, the stability also remarkably increased. The mucoadhesive capacity of the

CS-coated liposomes was much better than that of the noncoated ones. The cytotoxicity of the RFM-loaded CS-coated liposomes on A549 cells was much lower than that of the free drug. The results showed that the CS coating could significantly enhance the mucoadhesive capacity of the liposomes, and these CS-coated liposomes could be potential drug carriers for pulmonary administration by nebulization.

protect the siRNA against the destructive shear forces generated by mesh-based nebulizers. Aerosol treatment also improved the size distribution of the NPs, which was beneficial in promoting the transfection efficiency. This CS-based targeting DDS had potential applications for siRNA delivery in lung disease treatment by

Suhui Ni et al. had prepared γ-aminobutyric acid type B (GABAB) receptor ligand-directed NPs for survivin siRNA delivery [83]. The NPs were synthesized by baclofen (Bac)-functionalized trimethyl CS (Bac-TMC) with tripolyphosphate (TPP) as an ionic crosslinker. GABAB receptor agonist Bac was initially introduced into TMC as a novel ligand. The Bac-TMC/TPP NPs could promote the cellular uptake of siRNA by interacting with the GABAB receptor, which further induced effective gene silence and cell apoptosis. Mannitol microparticles were further utilized for the pulmonary delivery of the siRNA-loaded NPs via pressurized metered dose inhalers (pMDI). The obtained formulation had good aerodynamic properties, which benefited the deep lung deposition. The pMDI formulation

Recently, Ting Wu et al. had developed a genipin-crosslinked carboxymethyl CS nanogel for lung-targeted delivery of isoniazid (INH) and rifampin (RMP) [84]. The dual drug-loaded nanogel particles (NGPs) had a uniform particle size from 60 to 130 nm with spherical morphology. The NGPs had sustained release behavior in simulated lung fluid. The dual drug-loaded NGPs had high antibacterial activity against multidrug-resistant Mycobacterium tuberculosis, and also could realize longterm antibacterial activity. Further in vivo evaluation exhibited that alveolar delivery of NGPs had satisfactory deposition of drug within the lung with lower toxicity. The results indicated that the NGPs would be a potential dosage form for treating

Pulmonary administration is a promising route for drug delivery to prevent and treat diseases, especially for delivering the drugs for lung diseases and some small molecule drugs with low absorption rate when in oral dosage form, also suitable for some traditional Chinese medicines and peptide or protein drugs. PDDS can effectively deliver the therapeutic drugs to the target sites, and improve the drug bioavailability and therapeutic efficacy. Inhalation is a noninvasive method for pulmonary administration, and the inhaled drugs can directly enter the blood circulation through the absorption of the alveolar epithelium. Pulmonary administration can enhance the drug absorption rate, reduce systemic side-effects, and improve the compliance of long-term medication. By transforming the drugs into dry powder inhalation formulations, drug degradation can be avoided. The therapeutic effect of the PDDS is mainly influenced by the physicochemical properties of the DDS, the dosage forms, and the administration devices, and also some other factors. The increase of the amount of drug delivered into the lung and the promotion of drug absorption rate are the key factors to improve the therapeutic efficiency of pulmonary administration. The application of sustained or controlled release preparations is an important method to prolong drug action time, enhance drug bioavailability, and improve patient compliance. In recent years, many controlled release preparations or active targeting preparations for pulmonary drug delivery have been constructed by drug-loaded microparticles, liposomes, and NPs. These multifunctional drug carriers have gained increasing popularity in pulmonary administration. It is really inspiring for us to see that some of the PDDS have already been applied to treat patients in clinic and become commercially available

containing Bac-TMC/TPP NPs could be a potential PDDS for siRNA.

against multidrug-resistant Mycobacterium tuberculosis.

4. Future perspectives

173

aerosol inhalation.

Applications of Chitosan in Pulmonary Drug Delivery DOI: http://dx.doi.org/10.5772/intechopen.87932

In another study, Mitsutaka Murata et al. investigated the surface modification of liposomes by CS for pulmonary delivery [74]. The surface of the liposomes was modified with CS oligosaccharide (oligoCS) and polyvinyl alcohol with a hydrophobic anchor (PVA-R). The association study showed that the PVA-R modification decreased the interaction between liposomes and A549 cells. By contrast, the oligoCS modification could significantly promote the cellular interaction. After pulmonary administration to rats, the peptide elcatonin (eCT) loaded oligoCS or PVA-R modified liposomes exhibited significantly enhanced therapeutic efficacy. The oligoCS-modified liposomes could adhere to the lung tissues and open the tight junctions between cells, which remarkably improved the drug absorption rate. On the other hand, the PVA-R-modified liposomes could induce a sustained absorption through the long-term eCT retention in lung fluid. The results verified that the surface-modification of liposomes with oligoCS and PVA-R could be used as effective PDDS for peptide pulmonary administration.

#### 3.2.4 CS-based active targeting DDS

The pulmonary inhalation preparations are usually directly transported into the lung. Thus, they can passively target to the lung tissues by pulmonary administration. The deposition site of the drugs in the lung can be controlled by regulating the physicochemical and functional properties of the drug carriers to meet the disease treatment requirements. In addition to passive targeting, active targeting systems have more applications in precise and efficient disease treatment. Active targeting systems utilize ligands or antibody-modified carriers to deliver the drugs to the specifically targeted tissues or cells, even intracellular organelles, thereby improving drug efficacy and reducing toxicity and side effects [75, 76]. The drug carriers could be chemically modified with active ligands, such as sugar moieties (galactose, mannose, and glucose) or specific ligands, such as folic acid (FA), transferrin (Tf), and Arg-Gly-Asp (RGD) peptide [77–79]. Antibody-mediated active targeting is also a primary strategy for delivering the drugs to the specific parts of the body [80].

Hu-Lin Jiang et al. had prepared a folate-CS-graft-polyethylenimine (FC-g-PEI) copolymer for cancer cell-targeting gene delivery [81]. FC-g-PEI could effectively load and protect the shRNA. The copolymer also showed decreased cytotoxicity compared with the PEI control. Compared with the untargeted polymer, FC-g-PEI was a more efficient Akt1 shRNA carrier, and it exhibited effective cancer celltargeting ability. Moreover, aerosol delivery of the FC-g-PEI/Akt1 shRNA NPs effectively inhibited lung tumorigenesis in the urethane-induced lung cancer model mice via Akt signaling pathway. The results demonstrated that the FC-g-PEI could be applied for the shRNA gene therapy via aerosol delivery.

Yongfeng Luo et al. developed an inhalable β2-adrenoceptor ligand-directed guanidinylated-CS (GCS) carrier for targeted lung delivery of siRNA [82]. GCS could effectively condense the siRNA and form complex NPs. Compared with pristine CS, the siRNA-loaded GCS NPs exhibited lower cytotoxicity and higher cellular internalization, which finally resulted in promoted gene silence efficiency. Salbutamol (a β2-adrenoceptor agonist) was further chemically coupled to the GCS to enhance the targeting specificity of the siRNA-loaded NPs. The gene silence efficacy was remarkably increased both in vitro and in vivo. The carrier could

Applications of Chitosan in Pulmonary Drug Delivery DOI: http://dx.doi.org/10.5772/intechopen.87932

CS-coated liposomes was much better than that of the noncoated ones. The cytotoxicity of the RFM-loaded CS-coated liposomes on A549 cells was much lower than that of the free drug. The results showed that the CS coating could significantly enhance the mucoadhesive capacity of the liposomes, and these CS-coated liposomes could be potential drug carriers for pulmonary administration by

In another study, Mitsutaka Murata et al. investigated the surface modification of liposomes by CS for pulmonary delivery [74]. The surface of the liposomes was modified with CS oligosaccharide (oligoCS) and polyvinyl alcohol with a hydrophobic anchor (PVA-R). The association study showed that the PVA-R modification decreased the interaction between liposomes and A549 cells. By contrast, the oligoCS modification could significantly promote the cellular interaction. After pulmonary administration to rats, the peptide elcatonin (eCT) loaded oligoCS or PVA-R modified liposomes exhibited significantly enhanced therapeutic efficacy. The oligoCS-modified liposomes could adhere to the lung tissues and open the tight junctions between cells, which remarkably improved the drug absorption rate. On the other hand, the PVA-R-modified liposomes could induce a sustained absorption through the long-term eCT retention in lung fluid. The results verified that the surface-modification of liposomes with oligoCS and PVA-R could be used as effec-

The pulmonary inhalation preparations are usually directly transported into the lung. Thus, they can passively target to the lung tissues by pulmonary administration. The deposition site of the drugs in the lung can be controlled by regulating the physicochemical and functional properties of the drug carriers to meet the disease treatment requirements. In addition to passive targeting, active targeting systems have more applications in precise and efficient disease treatment. Active targeting systems utilize ligands or antibody-modified carriers to deliver the drugs to the specifically targeted tissues or cells, even intracellular organelles, thereby improving drug efficacy and reducing toxicity and side effects [75, 76]. The drug carriers could be chemically modified with active ligands, such as sugar moieties (galactose, mannose, and glucose) or specific ligands, such as folic acid (FA), transferrin (Tf), and Arg-Gly-Asp (RGD) peptide [77–79]. Antibody-mediated active targeting is also a primary strategy for delivering the drugs to the specific parts of the body [80].

Hu-Lin Jiang et al. had prepared a folate-CS-graft-polyethylenimine (FC-g-PEI) copolymer for cancer cell-targeting gene delivery [81]. FC-g-PEI could effectively load and protect the shRNA. The copolymer also showed decreased cytotoxicity compared with the PEI control. Compared with the untargeted polymer, FC-g-PEI was a more efficient Akt1 shRNA carrier, and it exhibited effective cancer celltargeting ability. Moreover, aerosol delivery of the FC-g-PEI/Akt1 shRNA NPs effectively inhibited lung tumorigenesis in the urethane-induced lung cancer model mice via Akt signaling pathway. The results demonstrated that the FC-g-PEI could

Yongfeng Luo et al. developed an inhalable β2-adrenoceptor ligand-directed guanidinylated-CS (GCS) carrier for targeted lung delivery of siRNA [82]. GCS could effectively condense the siRNA and form complex NPs. Compared with pristine CS, the siRNA-loaded GCS NPs exhibited lower cytotoxicity and higher cellular internalization, which finally resulted in promoted gene silence efficiency. Salbutamol (a β2-adrenoceptor agonist) was further chemically coupled to the GCS to enhance the targeting specificity of the siRNA-loaded NPs. The gene silence efficacy was remarkably increased both in vitro and in vivo. The carrier could

be applied for the shRNA gene therapy via aerosol delivery.

tive PDDS for peptide pulmonary administration.

Role of Novel Drug Delivery Vehicles in Nanobiomedicine

3.2.4 CS-based active targeting DDS

nebulization.

172

protect the siRNA against the destructive shear forces generated by mesh-based nebulizers. Aerosol treatment also improved the size distribution of the NPs, which was beneficial in promoting the transfection efficiency. This CS-based targeting DDS had potential applications for siRNA delivery in lung disease treatment by aerosol inhalation.

Suhui Ni et al. had prepared γ-aminobutyric acid type B (GABAB) receptor ligand-directed NPs for survivin siRNA delivery [83]. The NPs were synthesized by baclofen (Bac)-functionalized trimethyl CS (Bac-TMC) with tripolyphosphate (TPP) as an ionic crosslinker. GABAB receptor agonist Bac was initially introduced into TMC as a novel ligand. The Bac-TMC/TPP NPs could promote the cellular uptake of siRNA by interacting with the GABAB receptor, which further induced effective gene silence and cell apoptosis. Mannitol microparticles were further utilized for the pulmonary delivery of the siRNA-loaded NPs via pressurized metered dose inhalers (pMDI). The obtained formulation had good aerodynamic properties, which benefited the deep lung deposition. The pMDI formulation containing Bac-TMC/TPP NPs could be a potential PDDS for siRNA.

Recently, Ting Wu et al. had developed a genipin-crosslinked carboxymethyl CS nanogel for lung-targeted delivery of isoniazid (INH) and rifampin (RMP) [84]. The dual drug-loaded nanogel particles (NGPs) had a uniform particle size from 60 to 130 nm with spherical morphology. The NGPs had sustained release behavior in simulated lung fluid. The dual drug-loaded NGPs had high antibacterial activity against multidrug-resistant Mycobacterium tuberculosis, and also could realize longterm antibacterial activity. Further in vivo evaluation exhibited that alveolar delivery of NGPs had satisfactory deposition of drug within the lung with lower toxicity. The results indicated that the NGPs would be a potential dosage form for treating against multidrug-resistant Mycobacterium tuberculosis.
