Effective nebulization of interferon-γ using a novel vibrating mesh

Abstract

Background: Interferon gamma (IFN-γ) is a clinically relevant immunomodulatory cytokine that has demonstrated significant potential in the treatment and management of respiratory diseases such as tuberculosis and pulmonary fibrosis. As with all large biomolecules, clinical translation is dependent on effective delivery to the disease site and delivery of IFN-γ as an aerosol offers a logical means of drug targeting. Effective localization is often hampered by instability and a lack of safe and efficient delivery systems. The present study sought to determine how effectively IFN-γ can be nebulized using two types of vibrating mesh nebulizer, each with differing mesh architectures, and to investigate the comparative efficiency of delivery of therapeutically active IFN-γ to the lungs.

Methods: Nebulization of IFN-γ was carried out using two different Aerogen vibrating mesh technologies with differing mesh architectures. These technologies represent both a standard commercially available mesh type (Aerogen Solo®) and a new iteration mesh (Photo-defined aperture plate (PDAP®). Extensive aerosol studies (aerosol output and droplet analysis, non-invasive and invasive aerosol therapy) were conducted in line with regulatory requirements and characterization of the stability and bioactivity of the IFN-γ post-nebulization was confirmed using SDS-PAGE and stimulation of Human C-X-C motif chemokine 10 (CXCL 10) also known as IFN-γ-induced protein 10KDa (IP 10) expression from THP-1 derived macrophages (THP-1 cells).

Results: Aerosol characterization studies indicated that a significant and reproducible dose of aerosolized IFN-γ can be delivered using both vibrating mesh technologies. Nebulization using both devices resulted in an emitted dose of at least 93% (100% dose minus residual volume) for IFN-γ. Characterization of aerosolized IFN-γ indicated that the PDAP was capable of generating droplets with a significantly lower mass median aerodynamic diameter (MMAD) with values of 2.79 ± 0.29 μm and 4.39 ± 0.25 μm for the PDAP and Solo respectively. The volume median diameters (VMD) of aerosolized IFN-γ corroborated this with VMDs of 2.33 ± 0.02 μm for the PDAP and 4.30 ± 0.02 μm for the Solo. SDS-PAGE gels indicated that IFN-γ remains stable after nebulization by both devices and this was confirmed by bioactivity studies using a THP-1 cell model in which an alveolar macrophage response to IFN-γ was determined. IFN-γ nebulized by the PDAP and Solo devices had no significant effect on the key inflammatory biomarker cytokine IP-10 release from this model in comparison to non-nebulized controls. Here we demonstrate that it is possible to combine IFN-γ with vibrating mesh nebulizer devices and facilitate effective aerosolisation with minimal impact on IFN-γ structure or bioactivity.

Conclusions: It is possible to nebulize IFN-γ effectively with vibrating mesh nebulizer devices without compromising its stability. The PDAP allows for generation of IFN-γ aerosols with improved aerodynamic properties thereby increasing its potential efficiency for lower respiratory tract deposition over current technology, whilst maintaining the integrity and bioactivity of IFN-γ. This delivery modality therefore offers a rational means of facilitating the clinical translation of inhaled IFN-γ.

Keywords: Aerosol; Idiopathic pulmonary fibrosis; Inhaled therapy; Interferon gamma; Nebulizer; Tuberculosis; Vibrating mesh.

Conflict of interest statement

Ethics approval and consent to participate

Not Applicable.

Consent for publication

Not Applicable.

Competing interests

Ronan MacLoughlin and Louise Sweeney are employees of Aerogen, the manufacturer of the vibrating mesh nebulizers described in this study.

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Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Figures

Fig. 1

Mesh architecture: a General perspective view of reservoir wells on a thick support layer overlaid on a thin outlet layer as captured by scanning electron microscope (SEM) (Magnification 817x). b Reservoir image of the PDAP (photo defined aperture plate) device as captured by SEM. c Outlet holes of the PDAP device aperture plate as captured by SEM. d Solo and PDAP mesh housing

Fig. 2

Tracheal Dose test setup with high flow nasal cannula, showing the adult head model attached to the breathing simulator

Fig. 3

IFN-γ recovery (pg) of 1 ml dose volumes of known doses of IFN-γ (500–125 pg/ml) using both VMNs (vibrating mesh nebulizer’s) - Solo and PDAP devices. Results were compared to non-nebulized controls (n = 6)

Fig. 4

SDS-PAGE analysis indicated that IFN-γ remained intact post-nebulization from both devices Aerogen Solo (lanes 4–6) and PDAP (lanes 7–9) in comparison to non-nebulized controls (lanes 2–3). The molecular weight marker is in lane 1

Fig. 5

CXCL-10 release from THP-1 cells (n = 5) following treatment with nebulized IFN-γ (using Solo and PDAP devices) and non-nebulized (control) IFN-γ at 10 and 5 ng/ml. Two additional controls were employed RPMI and IFN-γ diluent and were non-detectable (ND)