Self-Assembly Properties of an Amphiphilic Phosphate Ester Prodrug Designed for the Treatment of COVID-19.

PF-07304814 is a water-soluble phosphate ester prodrug of a small molecule inhibitor for the SARS CoV-2 3CL protease designed for the treatment of COVID-19. The amphiphilicity and self-assembly behavior of the prodrug was investigated computationally and experimentally via multiple orthogonal techniques to better design formulations for intravenous infusion. The self-assembly of PF-07304814 into micellar structures enabled an increase in the solubility of lipophilic impurities by up to 1900x in clinically relevant formulations. The observed solubilization could help extend the drug product shelf-life and in use stability through inhibition of precipitation, without the need for solubilizing excipients. The work presented in this manuscript provides a roadmap for the characterization of prodrug self-assembly and highlights the potential for prodrug modifications to enhance solubility of both active ingredients and impurities and to extend drug product shelf-life.

Preclinical characterization of an intravenous coronavirus 3CL protease inhibitor for the potential treatment of COVID19.

COVID-19 caused by the SARS-CoV-2 virus has become a global pandemic. 3CL protease is a virally encoded protein that is essential across a broad spectrum of coronaviruses with no close human analogs. PF-00835231, a 3CL protease inhibitor, has exhibited potent in vitro antiviral activity against SARS-CoV-2 as a single agent. Here we report, the design and characterization of a phosphate prodrug PF-07304814 to enable the delivery and projected sustained systemic exposure in human of PF-00835231 to inhibit coronavirus family 3CL protease activity with selectivity over human host protease targets. Furthermore, we show that PF-00835231 has additive/synergistic activity in combination with remdesivir. We present the ADME, safety, in vitro, and in vivo antiviral activity data that supports the clinical evaluation of PF-07304814 as a potential COVID-19 treatment.

Discovery of a Novel Inhibitor of Coronavirus 3CL Protease for the Potential Treatment of COVID-19.

COVID-19 caused by the SARS-CoV-2 virus has become a global pandemic. 3CL protease is a virally encoded protein that is essential across a broad spectrum of coronaviruses with no close human analogs. The designed phosphate prodrug PF-07304814 is metabolized to PF-00835321 which is a potent inhibitor in vitro of the coronavirus family 3CL pro, with selectivity over human host protease targets. Furthermore, PF-00835231 exhibits potent in vitro antiviral activity against SARS-CoV-2 as a single agent and it is additive/synergistic in combination with remdesivir. We present the ADME, safety, in vitro , and in vivo antiviral activity data that supports the clinical evaluation of this compound as a potential COVID-19 treatment.

Integration of active pharmaceutical ingredient solid form selection and particle engineering into drug product design.

This review seeks to offer a broad perspective that encompasses an understanding of the drug product attributes affected by active pharmaceutical ingredient (API) physical properties, their link to solid form selection and the role of particle engineering. While the crucial role of active pharmaceutical ingredient (API) solid form selection is universally acknowledged in the pharmaceutical industry, the value of increasing effort to understanding the link between solid form, API physical properties and drug product formulation and manufacture is now also being recognised. A truly holistic strategy for drug product development should focus on connecting solid form selection, particle engineering and formulation design to both exploit opportunities to access simpler manufacturing operations and prevent failures. Modelling and predictive tools that assist in establishing these links early in product development are discussed. In addition, the potential for differences between the ingoing API physical properties and those in the final product caused by drug product processing is considered. The focus of this review is on oral solid dosage forms and dry powder inhaler products for lung delivery.

The influence of physical properties and morphology of crystallised lactose on delivery of salbutamol sulphate from dry powder inhalers.

The aim of this work was to investigate the mechanistic evaluation of physicochemical properties of new engineered lactose on aerosolisation performance of salbutamol sulphate (SS) delivered from dry powder inhaler (DPI). Different crystallised lactose particles were obtained from binary mixtures of butanol:acetone. The sieved fractions (63-90 µm) of crystallised lactose were characterised in terms of size, shape, flowability, true density and aerosolisation performance (using multiple twin stage impinger (MSLI), Aerolizer(®) inhaler device, and salbutamol sulphate as a model drug). Compared to commercial lactose, crystallised lactose particles were less elongated, covered with fine lactose particles, and had a rougher surface morphology. The crystallised lactose powders had a considerably lower bulk and tap density and poorer flow when compared to commercial lactose. Engineered carrier with better flow showed improved drug content homogeneity, reduced amounts of drug "deposited" on the inhaler device and throat, and a smaller drug aerodynamic diameter upon inhalation. Aerodynamic diameter of salbutamol sulphate increased as lactose aerodynamic diameter decreased (linear, R(2)=0.9191) and/or as fine particle lactose content increased (linear, R(2)=0.8653). Improved drug aerosolisation performance in the case of crystallised lactose particles was attributed to lower drug-carrier adhesion forces due to a rougher surface and higher fine particle content. In conclusion, this work proved that using binary combinations of solvents in crystallisation medium is vital in modification of the physicochemical and micromeritic properties of carriers to achieve a desirable aerosolisation performance from DPI formulations. Among all lactose samples, lactose particles crystallised from pure butanol generated the highest overall DPI formulations desirability.

Dry powder inhalers: mechanistic evaluation of lactose formulations containing salbutamol sulphate.

The purpose of this study was to evaluate the relationships between physicochemical properties and aerosolisation performance of different grades of lactose. In order to get a wide range of physicochemical properties, various grades of lactose namely Flowlac 100 (FLO), Lactopress anhydrous 250 (LAC), Cellactose 80 (CEL), Tablettose 80 (TAB), and Granulac 200 (GRA) were used. The different lactose grades were carefully sieved to separate 63-90 µm particle size fractions and then characterised in terms of size, shape, density, flowability, and solid state. Formulations were prepared by blending each lactose with salbutamol sulphate (SS) at ratio of 67.5:1 (w/w), and then evaluated in terms of SS content uniformity, lactose-SS adhesion properties, and in vitro aerosolisation performance delivered from the Aerolizer. Sieved lactose grades showed similar particle size distributions (PSDs) and good flow properties but different particle shape, particle surface texture, and particle solid state. Content uniformity assessments indicated that lactose particles with rougher surface produced improved SS homogeneity within DPI formulation powders. Lactose-SS adhesion assessments indicated that lactose particles with more elongated shape and the rougher surface showed smaller adhesion force between lactose and salbutamol sulphate. Lactose powders with higher bulk density and higher tap density produced smaller emission (EM) and higher drug loss (DL) of SS. In vitro aerosolisation for various lactose grades followed the following rank order in terms of deposition performance: GRA>TAB>LAC ≈ CEL>FLO. Linear relationships were established showing that in order to maximize SS delivery to lower airway regions, lactose particles with more elongated shape, more irregular shape, and rougher surface are preferred. Therefore, considerable improvement in DPI performance can be achieved by careful selection of grade of lactose included within DPI formulations.

Improved aerosolization performance of salbutamol sulfate formulated with lactose crystallized from binary mixtures of ethanol-acetone.

It has been shown that dry powder inhaler (DPI) formulations typically achieve low fine particle fractions (poor performance). A commonly held theory is that this is due, at least in part, to low levels of detachment of drug from lactose during aerosolization as a result of strong adhesion of drug particles to the carrier surfaces. Therefore, the purpose of the present study is to overcome poor aerosolization performance of DPI formulation by modification of lactose particles. Lactose particles were crystallized by adding solution in water to different ratios of binary mixtures of ethanol-acetone. The results showed that modified lactose particles had exceptional aerosolization performance that makes them superior to commercial lactose particles. Morphology assessment showed that crystallized lactose particles were less elongated, more irregular in shape, and composed of smaller primary lactose particles compared with commercial lactose. Solid-state characterization showed that commercial lactose particles were α-lactose monohydrate, whereas crystallized lactose particles were a mixture of α-lactose monohydrate and ß-lactose according to the ratio of ethanol-acetone used during crystallization process. The enhanced performance could be mainly due to rougher surface and/or higher amounts of fines compared with the lactose crystallized from pure ethanol or commercial lactose.

Characterisation and deposition studies of recrystallised lactose from binary mixtures of ethanol/butanol for improved drug delivery from dry powder inhalers.

Dry powder inhaler formulations comprising commercial lactose-drug blends can show restricted detachment of drug from lactose during aerosolisation, which can lead to poor fine particle fractions (FPFs) which are suboptimal. The aim of the present study was to investigate whether the crystallisation of lactose from different ethanol/butanol co-solvent mixtures could be employed as a method of altering the FPF of salbutamol sulphate from powder blends. Lactose particles were prepared by an anti-solvent recrystallisation process using various ratios of the two solvents. Crystallised lactose or commercial lactose was mixed with salbutamol sulphate and in vitro deposition studies were performed using a multistage liquid impinger. Solid-state characterisation results showed that commercial lactose was primarily composed of the α-anomer whilst the crystallised lactose samples comprised a α/ß mixture containing a lower number of moles of water per mole of lactose compared to the commercial lactose. The crystallised lactose particles were also less elongated and more irregular in shape with rougher surfaces. Formulation blends containing crystallised lactose showed better aerosolisation performance and dose uniformity when compared to commercial lactose. The highest FPF of salbutamol sulphate (38.0 ± 2.5%) was obtained for the lactose samples that were crystallised from a mixture of ethanol/butanol (20:60) compared to a FPF of 19.7 ± 1.9% obtained for commercial lactose. Engineered lactose carriers with modified anomer content and physicochemical properties, when compared to the commercial grade, produced formulations which generated a high FPF.

Engineered mannitol as an alternative carrier to enhance deep lung penetration of salbutamol sulphate from dry powder inhaler.

In this research mannitol particles were prepared by recrystallisation using non-solvent precipitation technique to investigate the effect of engineered carrier particles on their physicochemical properties and the in vitro deposition profiles of a model drug (salbutamol sulphate (SS)) from a dry powder inhaler (DPI). To this end, mannitol aqueous solution (15%, w/v) was added to different ratios of ethanol:water (100:0, 95:5, 90:10 and 85:15) to obtain mannitol particles. These crystallised mannitol particles were analysed in terms of micromeritic properties, morphology, DSC, FT-IR, and in vitro fine particle fraction (FPF) and emitted dose (ED) of SS. The results showed that the elongation ratio of all the recrystallised mannitol batches was higher than the original material giving them a needle-shaped morphology. Salbutamol sulphate deposition profiles from DPI formulation containing recrystallised needle-shaped mannitol showed enhanced performance and better delivery to the lower MSLI stages. The FPF increased from 15.4+/-1.1 to 45.8+/-0.7% when the commercial mannitol was replaced by mannitol crystallised from ethanol:water (90:10). This improvement could be due to the presence of elongated mannitol crystals in formulation blends. Solid state characterisation of engineered mannitol showed that the commercial mannitol was beta-form, mannitol recrystallised from ethanol:water (85:15) was alpha-form and that samples recrystallised in presence of pure ethanol or other ratios of ethanol:water (95:5 and 90:10) were the mixtures of alpha-, beta- and delta-forms. Multi-solvent recrystallisation technique was proved to have potential to produce mannitol crystals suitable for enhanced aerosolisation efficiency. Comparing different crystallised mannitol formulations showed that the final form (the type of polymorph) of the crystallised mannitol does not have a substantial effect on salbutamol sulphate aerosolisation performance.

The enhanced aerosol performance of salbutamol from dry powders containing engineered mannitol as excipient.

The aim of the present study was to investigate the effect of crystallising mannitol from different binary mixtures of acetone/water on the resultant physical properties and to determine the effects of any changes on in vitro aerosolisation performance, when the different mannitol crystals were used as a carrier in dry powder inhaler formulations containing salbutamol sulphate. Mannitol particles were crystallised under controlled conditions by dissolving the sugar in water and precipitating the sugar using binary mixtures of acetone/water in different percentages as anti-solvent media. For comparison purposes the physical properties and deposition behaviour of commercially available mannitol were also studied. SEM showed that all crystallised mannitol particles were more elongated than the commercial mannitol. Solid state studies revealed that commercial mannitol and mannitol crystallised using acetone in the presence of 10-25% v/v water as anti-solvent was beta-polymorphic form whereas mannitol crystallised in the presence of a small amount of water (0-7.5%) was the alpha-form. All the crystallised mannitol samples showed poor flowability. Nevertheless, the powdered crystallised mannitol and commercial samples were blended with salbutamol in the ratio 67.5:1. The aerosolisation performance of the formulations containing the engineered mannitol (evaluated using Multi Stage Liquid Impinger) was considerably better than that of the commercial mannitol formulation (the fine particle fraction was increased from 15.42% to 33.07-43.99%, for the formulations containing crystallised mannitol). Generally, carriers having a high tapped density and high fraction of fine carrier particles produced a high FPF. The improvement in the DPI performance could be attributed to the presence of elongated carrier particles with smooth surfaces since these are believed to have less adhesive forces between carrier and the drug resulting in easier detachment of the drug during the inhalation.

Application of slurry bridging experiments at controlled water activities to predict the solid-state conversion between anhydrous and hydrated forms using theophylline as a model drug.

The role of water activity (a(w)), relative humidity (RH) and temperature on the hydration state of theophylline has been investigated. Slurry bridging experiments at controlled water activities, using powder X-ray diffraction (PXRD) and thermogravimetric analysis (TGA) to characterise the solid phase, established that the hydrate is the thermodynamically stable form of theophylline at a(w) > or = 0.5 at 4 degrees C, a(w) > or = 0.64 at 30 degrees C, and a(w) > or = 0.76 at 40 degrees C. These data were used to produce a phase stability diagram for anhydrous/hydrate theophylline versus temperature. Anhydrous theophylline was spray dried in an attempt to reduce crystallinity. The spray dried theophylline was stored at a range of temperatures (4-40 degrees C) and humidities (22-89% RH). Samples were analysed at 3, 6, 9, 26 and 52 weeks using TGA and at the 26 and 52 weeks by PXRD. The solid state stability of the spray dried theophylline closely correlated to the phase stability diagram produced using the slurry bridging experiments. The data suggest that the slurry bridging technique at controlled water activities provides an accurate method of rapidly predicting the physically stable form in anhydrous/hydrate systems.