| Popis: |
CHAPTER 1 This chapter comprises a literature review exploring the potential role of inhaled anti-inflammatory drugs as an effective treatment option for cystic fibrosis (CF) patients, in comparison with current oral delivery is presented. Specifically, the focus is on ibuprofen (IBU), the only non-steroidal anti-inflammatory drug approved for chronic use in CF. The need for inhalation therapy and the various delivery platforms for inhalation has also been highlighted with an insight into the reasons and challenges associated with developing an inhalation therapy of non-steroidal anti-inflammatory drugs (NSAIDs). This chapter has been published in Expert Opinion on Orphan Drugs, 2018. 6 (1): 69 - 84 under the title “Is there a role for inhaled anti-inflammatory drugs in cystic fibrosis treatment?” Authors: Z. Sheikh, H.X. Ong, M. Pozzoli, P.M. Young, D. Traini CHAPTER 2 This chapter focused on understanding the mechanisms of interactions between the anti-inflammatory drug ibuprofen (IBU) with each of the CF-approved inhaled antibiotics (Tobramycin/TOBI, Colistin/COL or the Colistin prodrug Tadim), including anti-bacterial and anti-inflammatory effects. Chemical interactions of the IBU: antibiotic combinations were elucidated using High-Resolution Mass-Spectrometry (HRMS) and 1H NMR. HRMS showed pairing of IBU and TOBI, further confirmed by 1H NMR, whilst no pairing was observed for either IBU: COL or IBU: Tadim. The anti-bacterial activity of the combinations against Pseudomonas aeruginosa (PAO) showed that neither paired nor non-paired IBU: antibiotic therapies altered the anti-bacterial activity. The anti-inflammatory efficacy of the combination therapies was then determined at two different concentrations (Low and High) using in vitro models of NuLi-1 (healthy) and CuFi-1 (CF) cell lines. Differential response in the anti-inflammatory efficacy of IBU: TOBI was observed between the two concentrations due to changes in the structural conformation of the paired IBU: TOBI complex at High concentration, confirmed by 1H NMR. In contrast, the non-pairing of the IBU: COL and IBU: Tadim combinations showed a significant decrease in IL-8 secretion at both the concentrations tested. Importantly, all antibiotics alone showed anti-inflammatory properties, highlighting the inherent anti-inflammatory properties of these antibiotics. These findings have significant implications for the delivery of combination treatments of IBU and the antibiotics and possible drug-drug interactions at the cellular level. This chapter is currently under review in European Journal of Pharmacology, 2020 under the title “Tobramycin and Colistin display anti-inflammatory properties in CuFi-1 cystic fibrosis cell line’’. Authors: Z. Sheikh, P. Bradbury, T.A. Reekie, M. Pozzoli, P.D. Robinson, M. Kassiou, P.M. Young, H.X. Ong, D. Traini CHAPTER 3 Following on from Chapter 2, Chapter 3 was aimed at developing an in vitro model with characteristic features of the CF airways for assessing drug transport of inhaled therapies used in CF treatment. Novel inhalable drug therapies require an in vitro CF model that appropriately mimics the in vivo CF lung environment to better understand drug delivery and transport across the CF epithelium, and predict drug therapeutic efficacy. In this context, an air-liquid interface (ALI) culture method of the CuFi-1 (CF cell line) was optimised and compared to the NuLi-1 (healthy cell line) cells concerning media composition, culture period and epithelial differentiation. Furthermore, drug transport on both CuFi-1 and NuLi-1 was investigated to determine whether these cell lines could be used to study the transport of drugs in CF treatment, using Ibuprofen as a model drug. Differentiating characteristics specific to airway epithelia such as mucus production, inflammatory response and tight junction formation at two seeding densities (Low and High) were assessed throughout an 8-week ALI culture period. This study demonstrated that both the NuLi-1 and CuFi-1 cell lines fully differentiate in ALI culture with significant mucus secretion, pro-inflammatory cytokines IL-6 and IL-8 production, and functional tight junctions at week 8. For the first time, this study identified that ibuprofen is transported via the paracellular pathway in ALI models of NuLi-1 and CuFi-1 cell lines. Overall, these findings highlighted NuLi-1 and CuFi-1 as promising in vitro ALI models to investigate the transport properties of novel inhalable drug therapies for CF treatment. This chapter has been published in European Journal of Pharmaceutics and Biopharmaceutics, 2020. 156: 121 – 130 under the title “An in vitro model for assessing drug transport in cystic fibrosis treatment: Characterisation of the CuFi-1 cell line”. Authors: Z. Sheikh, P. Bradbury, M. Pozzoli, P.M. Young, H.X. Ong, D. Traini CHAPTER 4 Once established that the ALI models of NuLi-1 and CuFi-1 are suitable cell culture models to study drug transport of inhaled therapies in Chapter 3, Chapter 4 focused on exploring diclofenac (DICLO) as an anti-inflammatory option for CF treatment using these in vitro ALI models of NuLi-1 and CuFi-1. As anti-inflammatory treatment options for CF are limited, there is a need to develop new anti-inflammatory agents. Therefore, a novel pressurized metered dose inhalation (pMDI) formulation of DICLO was successfully developed with the subsequent characterization of the in vitro aerosol performance, transport properties and anti-inflammatory activity across the NuLi-1 and CuFi-1 ALI models. A significantly higher amount of DICLO was transported across the ALI cultures for simulated inhaled delivery in comparison to simulated oral delivery, suggesting the potential of using the inhaled route for targeted delivery of DICLO to the lungs. Consequently, two DICLO pMDI formulations were developed to allow targeted DICLO delivery to CF airways: a Low dose formulation (delivering 150 µg of DICLO per dose) and a High dose formulation (delivering 430 µg of DICLO per dose) and characterized to determine the optimum formulation. The Low dose pMDI DICLO formulation showed a significantly smaller particle size diameter with uniform distribution resulting in a greater aerosol performance when compared to High dose formulation. Importantly, the Low dose DICLO pMDI was efficiently transported across the ALI models and displayed anti-inflammatory activity in both healthy and CF in vitro models, overall highlighting the potential of an aerosolized low-dose DICLO formulation as a promising inhaled anti-inflammatory therapy for CF treatment. This chapter is currently under review for publication in International Journal of Pharmaceutics, 2020 with the title “Development and in vitro characterization of a novel pMDI diclofenac formulation as an inhalable anti-inflammatory therapy for cystic fibrosis”. Authors: Z. Sheikh, L.G. Dos Reis, P. Bradbury, G. Meneguzzo, S. Scalia, P.M. Young, H.X. Ong, D. Traini CHAPTER 5 Ultimately, Chapter 5 summarizes the overall findings presented in this thesis including prospects. The inhaled route allows targeted drug delivery to the lungs at low doses, overcoming decreased bioavailability issues and other systemic side-effects associated with high oral doses. Owing to these numerous advantages of inhaled delivery, the role of potential inhaled anti-inflammatory drugs used in CF treatment with a particular focus on ibuprofen has been investigated. The mechanisms of interactions between the two primary modalities of CF treatment – anti-inflammatory drugs (ibuprofen) and antibiotics was further elucidated to determine if a combination therapy could be more beneficial. Surprisingly, ibuprofen in combination with the antibiotics did not significantly decrease inflammation in a CF model when compared to the drugs alone, suggesting that new anti-inflammatory drugs must be explored to identify a more effective combination treatment. However, to study the drug delivery of novel therapies, an in vitro model simulating the CF lung and suitable to assess drug transport is required. Hence, an in vitro CF bronchial epithelium model to study drug transport was developed and characterized to recapitulate the CF airways and compared to a healthy epithelial model. Finally, a low dose inhalation formulation of novel anti-inflammatory drug diclofenac was successfully developed and characterized in terms of transport and anti-inflammatory activity using these established models. Future studies will focus on investigating the interactions of diclofenac with the CF-approved antibiotics on the developed models and correlate these findings in vivo to comprehend its clinical relevance to optimize CF treatment and reduce treatment burden of CF patients. |
